Links to Exploration step
Le document en format XML
<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Structural Definition of a Neutralization-Sensitive Epitope on the MERS-CoV S1-NTD</title><author><name sortKey="Wang, Nianshuang" sort="Wang, Nianshuang" uniqKey="Wang N" first="Nianshuang" last="Wang">Nianshuang Wang</name><affiliation><nlm:aff id="aff1">Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX 78712, USA</nlm:aff></affiliation></author><author><name sortKey="Rosen, Osnat" sort="Rosen, Osnat" uniqKey="Rosen O" first="Osnat" last="Rosen">Osnat Rosen</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Wang, Lingshu" sort="Wang, Lingshu" uniqKey="Wang L" first="Lingshu" last="Wang">Lingshu Wang</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Turner, Hannah L" sort="Turner, Hannah L" uniqKey="Turner H" first="Hannah L." last="Turner">Hannah L. Turner</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Stevens, Laura J" sort="Stevens, Laura J" uniqKey="Stevens L" first="Laura J." last="Stevens">Laura J. Stevens</name><affiliation><nlm:aff id="aff4">Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation></author><author><name sortKey="Corbett, Kizzmekia S" sort="Corbett, Kizzmekia S" uniqKey="Corbett K" first="Kizzmekia S." last="Corbett">Kizzmekia S. Corbett</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Bowman, Charles A" sort="Bowman, Charles A" uniqKey="Bowman C" first="Charles A." last="Bowman">Charles A. Bowman</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Pallesen, Jesper" sort="Pallesen, Jesper" uniqKey="Pallesen J" first="Jesper" last="Pallesen">Jesper Pallesen</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Shi, Wei" sort="Shi, Wei" uniqKey="Shi W" first="Wei" last="Shi">Wei Shi</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Zhang, Yi" sort="Zhang, Yi" uniqKey="Zhang Y" first="Yi" last="Zhang">Yi Zhang</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Leung, Kwanyee" sort="Leung, Kwanyee" uniqKey="Leung K" first="Kwanyee" last="Leung">Kwanyee Leung</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Kirchdoerfer, Robert N" sort="Kirchdoerfer, Robert N" uniqKey="Kirchdoerfer R" first="Robert N." last="Kirchdoerfer">Robert N. Kirchdoerfer</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Becker, Michelle M" sort="Becker, Michelle M" uniqKey="Becker M" first="Michelle M." last="Becker">Michelle M. Becker</name><affiliation><nlm:aff id="aff4">Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation></author><author><name sortKey="Denison, Mark R" sort="Denison, Mark R" uniqKey="Denison M" first="Mark R." last="Denison">Mark R. Denison</name><affiliation><nlm:aff id="aff4">Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation><affiliation><nlm:aff id="aff5">Department of Pathology, Microbiology, and Immunology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation></author><author><name sortKey="Chappell, James D" sort="Chappell, James D" uniqKey="Chappell J" first="James D." last="Chappell">James D. Chappell</name><affiliation><nlm:aff id="aff4">Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation></author><author><name sortKey="Ward, Andrew B" sort="Ward, Andrew B" uniqKey="Ward A" first="Andrew B." last="Ward">Andrew B. Ward</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Graham, Barney S" sort="Graham, Barney S" uniqKey="Graham B" first="Barney S." last="Graham">Barney S. Graham</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Mclellan, Jason S" sort="Mclellan, Jason S" uniqKey="Mclellan J" first="Jason S." last="Mclellan">Jason S. Mclellan</name><affiliation><nlm:aff id="aff1">Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX 78712, USA</nlm:aff></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">31553909</idno><idno type="pmc">6935267</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6935267</idno><idno type="RBID">PMC:6935267</idno><idno type="doi">10.1016/j.celrep.2019.08.052</idno><date when="2019">2019</date><idno type="wicri:Area/Pmc/Corpus">000B58</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000B58</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Structural Definition of a Neutralization-Sensitive Epitope on the MERS-CoV S1-NTD</title><author><name sortKey="Wang, Nianshuang" sort="Wang, Nianshuang" uniqKey="Wang N" first="Nianshuang" last="Wang">Nianshuang Wang</name><affiliation><nlm:aff id="aff1">Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX 78712, USA</nlm:aff></affiliation></author><author><name sortKey="Rosen, Osnat" sort="Rosen, Osnat" uniqKey="Rosen O" first="Osnat" last="Rosen">Osnat Rosen</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Wang, Lingshu" sort="Wang, Lingshu" uniqKey="Wang L" first="Lingshu" last="Wang">Lingshu Wang</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Turner, Hannah L" sort="Turner, Hannah L" uniqKey="Turner H" first="Hannah L." last="Turner">Hannah L. Turner</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Stevens, Laura J" sort="Stevens, Laura J" uniqKey="Stevens L" first="Laura J." last="Stevens">Laura J. Stevens</name><affiliation><nlm:aff id="aff4">Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation></author><author><name sortKey="Corbett, Kizzmekia S" sort="Corbett, Kizzmekia S" uniqKey="Corbett K" first="Kizzmekia S." last="Corbett">Kizzmekia S. Corbett</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Bowman, Charles A" sort="Bowman, Charles A" uniqKey="Bowman C" first="Charles A." last="Bowman">Charles A. Bowman</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Pallesen, Jesper" sort="Pallesen, Jesper" uniqKey="Pallesen J" first="Jesper" last="Pallesen">Jesper Pallesen</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Shi, Wei" sort="Shi, Wei" uniqKey="Shi W" first="Wei" last="Shi">Wei Shi</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Zhang, Yi" sort="Zhang, Yi" uniqKey="Zhang Y" first="Yi" last="Zhang">Yi Zhang</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Leung, Kwanyee" sort="Leung, Kwanyee" uniqKey="Leung K" first="Kwanyee" last="Leung">Kwanyee Leung</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Kirchdoerfer, Robert N" sort="Kirchdoerfer, Robert N" uniqKey="Kirchdoerfer R" first="Robert N." last="Kirchdoerfer">Robert N. Kirchdoerfer</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Becker, Michelle M" sort="Becker, Michelle M" uniqKey="Becker M" first="Michelle M." last="Becker">Michelle M. Becker</name><affiliation><nlm:aff id="aff4">Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation></author><author><name sortKey="Denison, Mark R" sort="Denison, Mark R" uniqKey="Denison M" first="Mark R." last="Denison">Mark R. Denison</name><affiliation><nlm:aff id="aff4">Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation><affiliation><nlm:aff id="aff5">Department of Pathology, Microbiology, and Immunology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation></author><author><name sortKey="Chappell, James D" sort="Chappell, James D" uniqKey="Chappell J" first="James D." last="Chappell">James D. Chappell</name><affiliation><nlm:aff id="aff4">Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</nlm:aff></affiliation></author><author><name sortKey="Ward, Andrew B" sort="Ward, Andrew B" uniqKey="Ward A" first="Andrew B." last="Ward">Andrew B. Ward</name><affiliation><nlm:aff id="aff3">Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</nlm:aff></affiliation></author><author><name sortKey="Graham, Barney S" sort="Graham, Barney S" uniqKey="Graham B" first="Barney S." last="Graham">Barney S. Graham</name><affiliation><nlm:aff id="aff2">Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</nlm:aff></affiliation></author><author><name sortKey="Mclellan, Jason S" sort="Mclellan, Jason S" uniqKey="Mclellan J" first="Jason S." last="Mclellan">Jason S. Mclellan</name><affiliation><nlm:aff id="aff1">Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX 78712, USA</nlm:aff></affiliation></author></analytic><series><title level="j">Cell Reports</title><idno type="eISSN">2211-1247</idno><imprint><date when="2019">2019</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><title>Summary</title><p>Middle East respiratory syndrome coronavirus (MERS-CoV) emerged into the human population in 2012 and has caused substantial morbidity and mortality. Potently neutralizing antibodies targeting the receptor-binding domain (RBD) on MERS-CoV spike (S) protein have been characterized, but much less is known about antibodies targeting non-RBD epitopes. Here, we report the structural and functional characterization of G2, a neutralizing antibody targeting the MERS-CoV S1 N-terminal domain (S1-NTD). Structures of G2 alone and in complex with the MERS-CoV S1-NTD define a site of vulnerability comprising two loops, each of which contain a residue mutated in G2-escape variants. Cell-surface binding studies and <italic>in vitro</italic> competition experiments demonstrate that G2 strongly disrupts the attachment of MERS-CoV S to its receptor, dipeptidyl peptidase-4 (DPP4), with the inhibition requiring the native trimeric S conformation. These results advance our understanding of antibody-mediated neutralization of coronaviruses and should facilitate the development of immunotherapeutics and vaccines against MERS-CoV.</p></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Adams, P D" uniqKey="Adams P">P.D. Adams</name></author><author><name sortKey="Grosse Kunstleve, R W" uniqKey="Grosse Kunstleve R">R.W. Grosse-Kunstleve</name></author><author><name sortKey="Hung, L W" uniqKey="Hung L">L.W. Hung</name></author><author><name sortKey="Ioerger, T R" uniqKey="Ioerger T">T.R. Ioerger</name></author><author><name sortKey="Mccoy, A J" uniqKey="Mccoy A">A.J. McCoy</name></author><author><name sortKey="Moriarty, N W" uniqKey="Moriarty N">N.W. Moriarty</name></author><author><name sortKey="Read, R J" uniqKey="Read R">R.J. Read</name></author><author><name sortKey="Sacchettini, J C" uniqKey="Sacchettini J">J.C. Sacchettini</name></author><author><name sortKey="Sauter, N K" uniqKey="Sauter N">N.K. Sauter</name></author><author><name sortKey="Terwilliger, T C" uniqKey="Terwilliger T">T.C. Terwilliger</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Almazan, F" uniqKey="Almazan F">F. Almazán</name></author><author><name sortKey="Dediego, M L" uniqKey="Dediego M">M.L. DeDiego</name></author><author><name sortKey="Sola, I" uniqKey="Sola I">I. Sola</name></author><author><name sortKey="Zu Iga, S" uniqKey="Zu Iga S">S. Zuñiga</name></author><author><name sortKey="Nieto Torres, J L" uniqKey="Nieto Torres J">J.L. Nieto-Torres</name></author><author><name sortKey="Marquez Jurado, S" uniqKey="Marquez Jurado S">S. Marquez-Jurado</name></author><author><name sortKey="Andres, G" uniqKey="Andres G">G. Andrés</name></author><author><name sortKey="Enjuanes, L" uniqKey="Enjuanes L">L. Enjuanes</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Assiri, A M" uniqKey="Assiri A">A.M. Assiri</name></author><author><name sortKey="Midgley, C M" uniqKey="Midgley C">C.M. Midgley</name></author><author><name sortKey="Abedi, G R" uniqKey="Abedi G">G.R. Abedi</name></author><author><name sortKey="Bin Saeed, A" uniqKey="Bin Saeed A">A. Bin Saeed</name></author><author><name sortKey="Almasri, M M" uniqKey="Almasri M">M.M. Almasri</name></author><author><name sortKey="Lu, X" uniqKey="Lu X">X. Lu</name></author><author><name sortKey="Al Abdely, H M" uniqKey="Al Abdely H">H.M. Al-Abdely</name></author><author><name sortKey="Abdalla, O" uniqKey="Abdalla O">O. Abdalla</name></author><author><name sortKey="Mohammed, M" uniqKey="Mohammed M">M. Mohammed</name></author><author><name sortKey="Algarni, H S" uniqKey="Algarni H">H.S. Algarni</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Azhar, E I" uniqKey="Azhar E">E.I. Azhar</name></author><author><name sortKey="El Kafrawy, S A" uniqKey="El Kafrawy S">S.A. El-Kafrawy</name></author><author><name sortKey="Farraj, S A" uniqKey="Farraj S">S.A. Farraj</name></author><author><name sortKey="Hassan, A M" uniqKey="Hassan A">A.M. Hassan</name></author><author><name sortKey="Al Saeed, M S" uniqKey="Al Saeed M">M.S. Al-Saeed</name></author><author><name sortKey="Hashem, A M" uniqKey="Hashem A">A.M. Hashem</name></author><author><name sortKey="Madani, T A" uniqKey="Madani T">T.A. Madani</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Battye, T G" uniqKey="Battye T">T.G. Battye</name></author><author><name sortKey="Kontogiannis, L" uniqKey="Kontogiannis L">L. Kontogiannis</name></author><author><name sortKey="Johnson, O" uniqKey="Johnson O">O. Johnson</name></author><author><name sortKey="Powell, H R" uniqKey="Powell H">H.R. Powell</name></author><author><name sortKey="Leslie, A G" uniqKey="Leslie A">A.G. Leslie</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Chan, C M" uniqKey="Chan C">C.M. Chan</name></author><author><name sortKey="Chu, H" uniqKey="Chu H">H. Chu</name></author><author><name sortKey="Wang, Y" uniqKey="Wang Y">Y. Wang</name></author><author><name sortKey="Wong, B H" uniqKey="Wong B">B.H. Wong</name></author><author><name sortKey="Zhao, X" uniqKey="Zhao X">X. Zhao</name></author><author><name sortKey="Zhou, J" uniqKey="Zhou J">J. Zhou</name></author><author><name sortKey="Yang, D" uniqKey="Yang D">D. Yang</name></author><author><name sortKey="Leung, S P" uniqKey="Leung S">S.P. Leung</name></author><author><name sortKey="Chan, J F" uniqKey="Chan J">J.F. Chan</name></author><author><name sortKey="Yeung, M L" uniqKey="Yeung M">M.L. Yeung</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Chen, Y" uniqKey="Chen Y">Y. Chen</name></author><author><name sortKey="Lu, S" uniqKey="Lu S">S. Lu</name></author><author><name sortKey="Jia, H" uniqKey="Jia H">H. Jia</name></author><author><name sortKey="Deng, Y" uniqKey="Deng Y">Y. Deng</name></author><author><name sortKey="Zhou, J" uniqKey="Zhou J">J. Zhou</name></author><author><name sortKey="Huang, B" uniqKey="Huang B">B. Huang</name></author><author><name sortKey="Yu, Y" uniqKey="Yu Y">Y. Yu</name></author><author><name sortKey="Lan, J" uniqKey="Lan J">J. Lan</name></author><author><name sortKey="Wang, W" uniqKey="Wang W">W. Wang</name></author><author><name sortKey="Lou, Y" uniqKey="Lou Y">Y. Lou</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Chu, H" uniqKey="Chu H">H. Chu</name></author><author><name sortKey="Chan, C M" uniqKey="Chan C">C.M. Chan</name></author><author><name sortKey="Zhang, X" uniqKey="Zhang X">X. Zhang</name></author><author><name sortKey="Wang, Y" uniqKey="Wang Y">Y. Wang</name></author><author><name sortKey="Yuan, S" uniqKey="Yuan S">S. Yuan</name></author><author><name sortKey="Zhou, J" uniqKey="Zhou J">J. Zhou</name></author><author><name sortKey="Au Yeung, R K" uniqKey="Au Yeung R">R.K. Au-Yeung</name></author><author><name sortKey="Sze, K H" uniqKey="Sze K">K.H. Sze</name></author><author><name sortKey="Yang, D" uniqKey="Yang D">D. Yang</name></author><author><name sortKey="Shuai, H" uniqKey="Shuai H">H. Shuai</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Conway, P" uniqKey="Conway P">P. Conway</name></author><author><name sortKey="Tyka, M D" uniqKey="Tyka M">M.D. Tyka</name></author><author><name sortKey="Dimaio, F" uniqKey="Dimaio F">F. DiMaio</name></author><author><name sortKey="Konerding, D E" uniqKey="Konerding D">D.E. Konerding</name></author><author><name sortKey="Baker, D" uniqKey="Baker D">D. Baker</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Corti, D" uniqKey="Corti D">D. Corti</name></author><author><name sortKey="Zhao, J" uniqKey="Zhao J">J. Zhao</name></author><author><name sortKey="Pedotti, M" uniqKey="Pedotti M">M. Pedotti</name></author><author><name sortKey="Simonelli, L" uniqKey="Simonelli L">L. Simonelli</name></author><author><name sortKey="Agnihothram, S" uniqKey="Agnihothram S">S. Agnihothram</name></author><author><name sortKey="Fett, C" uniqKey="Fett C">C. Fett</name></author><author><name sortKey="Fernandez Rodriguez, B" uniqKey="Fernandez Rodriguez B">B. Fernandez-Rodriguez</name></author><author><name sortKey="Foglierini, M" uniqKey="Foglierini M">M. Foglierini</name></author><author><name sortKey="Agatic, G" uniqKey="Agatic G">G. Agatic</name></author><author><name sortKey="Vanzetta, F" uniqKey="Vanzetta F">F. Vanzetta</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Coughlin, M M" uniqKey="Coughlin M">M.M. Coughlin</name></author><author><name sortKey="Babcook, J" uniqKey="Babcook J">J. Babcook</name></author><author><name sortKey="Prabhakar, B S" uniqKey="Prabhakar B">B.S. Prabhakar</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Drosten, C" uniqKey="Drosten C">C. Drosten</name></author><author><name sortKey="Muth, D" uniqKey="Muth D">D. Muth</name></author><author><name sortKey="Corman, V M" uniqKey="Corman V">V.M. Corman</name></author><author><name sortKey="Hussain, R" uniqKey="Hussain R">R. Hussain</name></author><author><name sortKey="Al Masri, M" uniqKey="Al Masri M">M. Al Masri</name></author><author><name sortKey="Hajomar, W" uniqKey="Hajomar W">W. HajOmar</name></author><author><name sortKey="Landt, O" uniqKey="Landt O">O. Landt</name></author><author><name sortKey="Assiri, A" uniqKey="Assiri A">A. Assiri</name></author><author><name sortKey="Eckerle, I" uniqKey="Eckerle I">I. Eckerle</name></author><author><name sortKey="Al Shangiti, A" uniqKey="Al Shangiti A">A. Al Shangiti</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Du, L" uniqKey="Du L">L. Du</name></author><author><name sortKey="Tai, W" uniqKey="Tai W">W. Tai</name></author><author><name sortKey="Yang, Y" uniqKey="Yang Y">Y. Yang</name></author><author><name sortKey="Zhao, G" uniqKey="Zhao G">G. Zhao</name></author><author><name sortKey="Zhu, Q" uniqKey="Zhu Q">Q. Zhu</name></author><author><name sortKey="Sun, S" uniqKey="Sun S">S. Sun</name></author><author><name sortKey="Liu, C" uniqKey="Liu C">C. Liu</name></author><author><name sortKey="Tao, X" uniqKey="Tao X">X. Tao</name></author><author><name sortKey="Tseng, C K" uniqKey="Tseng C">C.K. Tseng</name></author><author><name sortKey="Perlman, S" uniqKey="Perlman S">S. Perlman</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Emsley, P" uniqKey="Emsley P">P. Emsley</name></author><author><name sortKey="Cowtan, K" uniqKey="Cowtan K">K. Cowtan</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Evans, P R" uniqKey="Evans P">P.R. Evans</name></author><author><name sortKey="Murshudov, G N" uniqKey="Murshudov G">G.N. Murshudov</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Fehr, A R" uniqKey="Fehr A">A.R. Fehr</name></author><author><name sortKey="Athmer, J" uniqKey="Athmer J">J. Athmer</name></author><author><name sortKey="Channappanavar, R" uniqKey="Channappanavar R">R. Channappanavar</name></author><author><name sortKey="Phillips, J M" uniqKey="Phillips J">J.M. Phillips</name></author><author><name sortKey="Meyerholz, D K" uniqKey="Meyerholz D">D.K. Meyerholz</name></author><author><name sortKey="Perlman, S" uniqKey="Perlman S">S. Perlman</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Gierer, S" uniqKey="Gierer S">S. Gierer</name></author><author><name sortKey="Bertram, S" uniqKey="Bertram S">S. Bertram</name></author><author><name sortKey="Kaup, F" uniqKey="Kaup F">F. Kaup</name></author><author><name sortKey="Wrensch, F" uniqKey="Wrensch F">F. Wrensch</name></author><author><name sortKey="Heurich, A" uniqKey="Heurich A">A. Heurich</name></author><author><name sortKey="Kr Mer Kuhl, A" uniqKey="Kr Mer Kuhl A">A. Krämer-Kühl</name></author><author><name sortKey="Welsch, K" uniqKey="Welsch K">K. Welsch</name></author><author><name sortKey="Winkler, M" uniqKey="Winkler M">M. Winkler</name></author><author><name sortKey="Meyer, B" uniqKey="Meyer B">B. Meyer</name></author><author><name sortKey="Drosten, C" uniqKey="Drosten C">C. Drosten</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Greenough, T C" uniqKey="Greenough T">T.C. Greenough</name></author><author><name sortKey="Babcock, G J" uniqKey="Babcock G">G.J. Babcock</name></author><author><name sortKey="Roberts, A" uniqKey="Roberts A">A. Roberts</name></author><author><name sortKey="Hernandez, H J" uniqKey="Hernandez H">H.J. Hernandez</name></author><author><name sortKey="Thomas, W D" uniqKey="Thomas W">W.D. Thomas</name></author><author><name sortKey="Coccia, J A" uniqKey="Coccia J">J.A. Coccia</name></author><author><name sortKey="Graziano, R F" uniqKey="Graziano R">R.F. Graziano</name></author><author><name sortKey="Srinivasan, M" uniqKey="Srinivasan M">M. Srinivasan</name></author><author><name sortKey="Lowy, I" uniqKey="Lowy I">I. Lowy</name></author><author><name sortKey="Finberg, R W" uniqKey="Finberg R">R.W. Finberg</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Gui, M" uniqKey="Gui M">M. Gui</name></author><author><name sortKey="Song, W" uniqKey="Song W">W. Song</name></author><author><name sortKey="Zhou, H" uniqKey="Zhou H">H. Zhou</name></author><author><name sortKey="Xu, J" uniqKey="Xu J">J. Xu</name></author><author><name sortKey="Chen, S" uniqKey="Chen S">S. Chen</name></author><author><name sortKey="Xiang, Y" uniqKey="Xiang Y">Y. Xiang</name></author><author><name sortKey="Wang, X" uniqKey="Wang X">X. Wang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Guo, Y" uniqKey="Guo Y">Y. Guo</name></author><author><name sortKey="Tisoncik, J" uniqKey="Tisoncik J">J. Tisoncik</name></author><author><name sortKey="Mcreynolds, S" uniqKey="Mcreynolds S">S. McReynolds</name></author><author><name sortKey="Farzan, M" uniqKey="Farzan M">M. Farzan</name></author><author><name sortKey="Prabhakar, B S" uniqKey="Prabhakar B">B.S. Prabhakar</name></author><author><name sortKey="Gallagher, T" uniqKey="Gallagher T">T. Gallagher</name></author><author><name sortKey="Rong, L" uniqKey="Rong L">L. Rong</name></author><author><name sortKey="Caffrey, M" uniqKey="Caffrey M">M. Caffrey</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Jiaming, L" uniqKey="Jiaming L">L. Jiaming</name></author><author><name sortKey="Yanfeng, Y" uniqKey="Yanfeng Y">Y. Yanfeng</name></author><author><name sortKey="Yao, D" uniqKey="Yao D">D. Yao</name></author><author><name sortKey="Yawei, H" uniqKey="Yawei H">H. Yawei</name></author><author><name sortKey="Linlin, B" uniqKey="Linlin B">B. Linlin</name></author><author><name sortKey="Baoying, H" uniqKey="Baoying H">H. Baoying</name></author><author><name sortKey="Jinghua, Y" uniqKey="Jinghua Y">Y. Jinghua</name></author><author><name sortKey="Gao, G F" uniqKey="Gao G">G.F. Gao</name></author><author><name sortKey="Chuan, Q" uniqKey="Chuan Q">Q. Chuan</name></author><author><name sortKey="Wenjie, T" uniqKey="Wenjie T">T. Wenjie</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Jiang, L" uniqKey="Jiang L">L. Jiang</name></author><author><name sortKey="Wang, N" uniqKey="Wang N">N. Wang</name></author><author><name sortKey="Zuo, T" uniqKey="Zuo T">T. Zuo</name></author><author><name sortKey="Shi, X" uniqKey="Shi X">X. Shi</name></author><author><name sortKey="Poon, K M" uniqKey="Poon K">K.M. Poon</name></author><author><name sortKey="Wu, Y" uniqKey="Wu Y">Y. Wu</name></author><author><name sortKey="Gao, F" uniqKey="Gao F">F. Gao</name></author><author><name sortKey="Li, D" uniqKey="Li D">D. Li</name></author><author><name sortKey="Wang, R" uniqKey="Wang R">R. Wang</name></author><author><name sortKey="Guo, J" uniqKey="Guo J">J. Guo</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Ki, M" uniqKey="Ki M">M. Ki</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Kirchdoerfer, R N" uniqKey="Kirchdoerfer R">R.N. Kirchdoerfer</name></author><author><name sortKey="Cottrell, C A" uniqKey="Cottrell C">C.A. Cottrell</name></author><author><name sortKey="Wang, N" uniqKey="Wang N">N. Wang</name></author><author><name sortKey="Pallesen, J" uniqKey="Pallesen J">J. Pallesen</name></author><author><name sortKey="Yassine, H M" uniqKey="Yassine H">H.M. Yassine</name></author><author><name sortKey="Turner, H L" uniqKey="Turner H">H.L. Turner</name></author><author><name sortKey="Corbett, K S" uniqKey="Corbett K">K.S. Corbett</name></author><author><name sortKey="Graham, B S" uniqKey="Graham B">B.S. Graham</name></author><author><name sortKey="Mclellan, J S" uniqKey="Mclellan J">J.S. McLellan</name></author><author><name sortKey="Ward, A B" uniqKey="Ward A">A.B. Ward</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Krempl, C" uniqKey="Krempl C">C. Krempl</name></author><author><name sortKey="Schultze, B" uniqKey="Schultze B">B. Schultze</name></author><author><name sortKey="Laude, H" uniqKey="Laude H">H. Laude</name></author><author><name sortKey="Herrler, G" uniqKey="Herrler G">G. Herrler</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Krissinel, E" uniqKey="Krissinel E">E. Krissinel</name></author><author><name sortKey="Henrick, K" uniqKey="Henrick K">K. Henrick</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Kunkel, F" uniqKey="Kunkel F">F. Künkel</name></author><author><name sortKey="Herrler, G" uniqKey="Herrler G">G. Herrler</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Lander, G C" uniqKey="Lander G">G.C. Lander</name></author><author><name sortKey="Stagg, S M" uniqKey="Stagg S">S.M. Stagg</name></author><author><name sortKey="Voss, N R" uniqKey="Voss N">N.R. Voss</name></author><author><name sortKey="Cheng, A" uniqKey="Cheng A">A. Cheng</name></author><author><name sortKey="Fellmann, D" uniqKey="Fellmann D">D. Fellmann</name></author><author><name sortKey="Pulokas, J" uniqKey="Pulokas J">J. Pulokas</name></author><author><name sortKey="Yoshioka, C" uniqKey="Yoshioka C">C. Yoshioka</name></author><author><name sortKey="Irving, C" uniqKey="Irving C">C. Irving</name></author><author><name sortKey="Mulder, A" uniqKey="Mulder A">A. Mulder</name></author><author><name sortKey="Lau, P W" uniqKey="Lau P">P.W. Lau</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Li, F" uniqKey="Li F">F. Li</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Li, F" uniqKey="Li F">F. Li</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Li, F" uniqKey="Li F">F. Li</name></author><author><name sortKey="Li, W" uniqKey="Li W">W. Li</name></author><author><name sortKey="Farzan, M" uniqKey="Farzan M">M. Farzan</name></author><author><name sortKey="Harrison, S C" uniqKey="Harrison S">S.C. Harrison</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Li, W" uniqKey="Li W">W. Li</name></author><author><name sortKey="Moore, M J" uniqKey="Moore M">M.J. Moore</name></author><author><name sortKey="Vasilieva, N" uniqKey="Vasilieva N">N. Vasilieva</name></author><author><name sortKey="Sui, J" uniqKey="Sui J">J. Sui</name></author><author><name sortKey="Wong, S K" uniqKey="Wong S">S.K. Wong</name></author><author><name sortKey="Berne, M A" uniqKey="Berne M">M.A. Berne</name></author><author><name sortKey="Somasundaran, M" uniqKey="Somasundaran M">M. Somasundaran</name></author><author><name sortKey="Sullivan, J L" uniqKey="Sullivan J">J.L. Sullivan</name></author><author><name sortKey="Luzuriaga, K" uniqKey="Luzuriaga K">K. Luzuriaga</name></author><author><name sortKey="Greenough, T C" uniqKey="Greenough T">T.C. Greenough</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Li, W" uniqKey="Li W">W. Li</name></author><author><name sortKey="Hulswit, R J G" uniqKey="Hulswit R">R.J.G. Hulswit</name></author><author><name sortKey="Widjaja, I" uniqKey="Widjaja I">I. Widjaja</name></author><author><name sortKey="Raj, V S" uniqKey="Raj V">V.S. Raj</name></author><author><name sortKey="Mcbride, R" uniqKey="Mcbride R">R. McBride</name></author><author><name sortKey="Peng, W" uniqKey="Peng W">W. Peng</name></author><author><name sortKey="Widagdo, W" uniqKey="Widagdo W">W. Widagdo</name></author><author><name sortKey="Tortorici, M A" uniqKey="Tortorici M">M.A. Tortorici</name></author><author><name sortKey="Van Dieren, B" uniqKey="Van Dieren B">B. van Dieren</name></author><author><name sortKey="Lang, Y" uniqKey="Lang Y">Y. Lang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Li, Y" uniqKey="Li Y">Y. Li</name></author><author><name sortKey="Wan, Y" uniqKey="Wan Y">Y. Wan</name></author><author><name sortKey="Liu, P" uniqKey="Liu P">P. Liu</name></author><author><name sortKey="Zhao, J" uniqKey="Zhao J">J. Zhao</name></author><author><name sortKey="Lu, G" uniqKey="Lu G">G. Lu</name></author><author><name sortKey="Qi, J" uniqKey="Qi J">J. Qi</name></author><author><name sortKey="Wang, Q" uniqKey="Wang Q">Q. Wang</name></author><author><name sortKey="Lu, X" uniqKey="Lu X">X. Lu</name></author><author><name sortKey="Wu, Y" uniqKey="Wu Y">Y. Wu</name></author><author><name sortKey="Liu, W" uniqKey="Liu W">W. Liu</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Lu, G" uniqKey="Lu G">G. Lu</name></author><author><name sortKey="Hu, Y" uniqKey="Hu Y">Y. Hu</name></author><author><name sortKey="Wang, Q" uniqKey="Wang Q">Q. Wang</name></author><author><name sortKey="Qi, J" uniqKey="Qi J">J. Qi</name></author><author><name sortKey="Gao, F" uniqKey="Gao F">F. Gao</name></author><author><name sortKey="Li, Y" uniqKey="Li Y">Y. Li</name></author><author><name sortKey="Zhang, Y" uniqKey="Zhang Y">Y. Zhang</name></author><author><name sortKey="Zhang, W" uniqKey="Zhang W">W. Zhang</name></author><author><name sortKey="Yuan, Y" uniqKey="Yuan Y">Y. Yuan</name></author><author><name sortKey="Bao, J" uniqKey="Bao J">J. Bao</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Mccoy, A J" uniqKey="Mccoy A">A.J. McCoy</name></author><author><name sortKey="Grosse Kunstleve, R W" uniqKey="Grosse Kunstleve R">R.W. Grosse-Kunstleve</name></author><author><name sortKey="Adams, P D" uniqKey="Adams P">P.D. Adams</name></author><author><name sortKey="Winn, M D" uniqKey="Winn M">M.D. Winn</name></author><author><name sortKey="Storoni, L C" uniqKey="Storoni L">L.C. Storoni</name></author><author><name sortKey="Read, R J" uniqKey="Read R">R.J. Read</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Millet, J K" uniqKey="Millet J">J.K. Millet</name></author><author><name sortKey="Whittaker, G R" uniqKey="Whittaker G">G.R. Whittaker</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Modjarrad, K" uniqKey="Modjarrad K">K. Modjarrad</name></author><author><name sortKey="Moorthy, V S" uniqKey="Moorthy V">V.S. Moorthy</name></author><author><name sortKey="Ben Embarek, P" uniqKey="Ben Embarek P">P. Ben Embarek</name></author><author><name sortKey="Van Kerkhove, M" uniqKey="Van Kerkhove M">M. Van Kerkhove</name></author><author><name sortKey="Kim, J" uniqKey="Kim J">J. Kim</name></author><author><name sortKey="Kieny, M P" uniqKey="Kieny M">M.P. Kieny</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Mohd, H A" uniqKey="Mohd H">H.A. Mohd</name></author><author><name sortKey="Al Tawfiq, J A" uniqKey="Al Tawfiq J">J.A. Al-Tawfiq</name></author><author><name sortKey="Memish, Z A" uniqKey="Memish Z">Z.A. Memish</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Nagae, M" uniqKey="Nagae M">M. Nagae</name></author><author><name sortKey="Ikeda, A" uniqKey="Ikeda A">A. Ikeda</name></author><author><name sortKey="Hane, M" uniqKey="Hane M">M. Hane</name></author><author><name sortKey="Hanashima, S" uniqKey="Hanashima S">S. Hanashima</name></author><author><name sortKey="Kitajima, K" uniqKey="Kitajima K">K. Kitajima</name></author><author><name sortKey="Sato, C" uniqKey="Sato C">C. Sato</name></author><author><name sortKey="Yamaguchi, Y" uniqKey="Yamaguchi Y">Y. Yamaguchi</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Naldini, L" uniqKey="Naldini L">L. Naldini</name></author><author><name sortKey="Blomer, U" uniqKey="Blomer U">U. Blömer</name></author><author><name sortKey="Gage, F H" uniqKey="Gage F">F.H. Gage</name></author><author><name sortKey="Trono, D" uniqKey="Trono D">D. Trono</name></author><author><name sortKey="Verma, I M" uniqKey="Verma I">I.M. Verma</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Oboho, I K" uniqKey="Oboho I">I.K. Oboho</name></author><author><name sortKey="Tomczyk, S M" uniqKey="Tomczyk S">S.M. Tomczyk</name></author><author><name sortKey="Al Asmari, A M" uniqKey="Al Asmari A">A.M. Al-Asmari</name></author><author><name sortKey="Banjar, A A" uniqKey="Banjar A">A.A. Banjar</name></author><author><name sortKey="Al Mugti, H" uniqKey="Al Mugti H">H. Al-Mugti</name></author><author><name sortKey="Aloraini, M S" uniqKey="Aloraini M">M.S. Aloraini</name></author><author><name sortKey="Alkhaldi, K Z" uniqKey="Alkhaldi K">K.Z. Alkhaldi</name></author><author><name sortKey="Almohammadi, E L" uniqKey="Almohammadi E">E.L. Almohammadi</name></author><author><name sortKey="Alraddadi, B M" uniqKey="Alraddadi B">B.M. Alraddadi</name></author><author><name sortKey="Gerber, S I" uniqKey="Gerber S">S.I. Gerber</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Ogura, T" uniqKey="Ogura T">T. Ogura</name></author><author><name sortKey="Iwasaki, K" uniqKey="Iwasaki K">K. Iwasaki</name></author><author><name sortKey="Sato, C" uniqKey="Sato C">C. Sato</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Pallesen, J" uniqKey="Pallesen J">J. Pallesen</name></author><author><name sortKey="Wang, N" uniqKey="Wang N">N. Wang</name></author><author><name sortKey="Corbett, K S" uniqKey="Corbett K">K.S. Corbett</name></author><author><name sortKey="Wrapp, D" uniqKey="Wrapp D">D. Wrapp</name></author><author><name sortKey="Kirchdoerfer, R N" uniqKey="Kirchdoerfer R">R.N. Kirchdoerfer</name></author><author><name sortKey="Turner, H L" uniqKey="Turner H">H.L. Turner</name></author><author><name sortKey="Cottrell, C A" uniqKey="Cottrell C">C.A. Cottrell</name></author><author><name sortKey="Becker, M M" uniqKey="Becker M">M.M. Becker</name></author><author><name sortKey="Wang, L" uniqKey="Wang L">L. Wang</name></author><author><name sortKey="Shi, W" uniqKey="Shi W">W. Shi</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Peng, G" uniqKey="Peng G">G. Peng</name></author><author><name sortKey="Sun, D" uniqKey="Sun D">D. Sun</name></author><author><name sortKey="Rajashankar, K R" uniqKey="Rajashankar K">K.R. Rajashankar</name></author><author><name sortKey="Qian, Z" uniqKey="Qian Z">Z. Qian</name></author><author><name sortKey="Holmes, K V" uniqKey="Holmes K">K.V. Holmes</name></author><author><name sortKey="Li, F" uniqKey="Li F">F. Li</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Peng, G" uniqKey="Peng G">G. Peng</name></author><author><name sortKey="Xu, L" uniqKey="Xu L">L. Xu</name></author><author><name sortKey="Lin, Y L" uniqKey="Lin Y">Y.L. Lin</name></author><author><name sortKey="Chen, L" uniqKey="Chen L">L. Chen</name></author><author><name sortKey="Pasquarella, J R" uniqKey="Pasquarella J">J.R. Pasquarella</name></author><author><name sortKey="Holmes, K V" uniqKey="Holmes K">K.V. Holmes</name></author><author><name sortKey="Li, F" uniqKey="Li F">F. Li</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Pettersen, E F" uniqKey="Pettersen E">E.F. Pettersen</name></author><author><name sortKey="Goddard, T D" uniqKey="Goddard T">T.D. Goddard</name></author><author><name sortKey="Huang, C C" uniqKey="Huang C">C.C. Huang</name></author><author><name sortKey="Couch, G S" uniqKey="Couch G">G.S. Couch</name></author><author><name sortKey="Greenblatt, D M" uniqKey="Greenblatt D">D.M. Greenblatt</name></author><author><name sortKey="Meng, E C" uniqKey="Meng E">E.C. Meng</name></author><author><name sortKey="Ferrin, T E" uniqKey="Ferrin T">T.E. Ferrin</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Potter, C S" uniqKey="Potter C">C.S. Potter</name></author><author><name sortKey="Chu, H" uniqKey="Chu H">H. Chu</name></author><author><name sortKey="Frey, B" uniqKey="Frey B">B. Frey</name></author><author><name sortKey="Green, C" uniqKey="Green C">C. Green</name></author><author><name sortKey="Kisseberth, N" uniqKey="Kisseberth N">N. Kisseberth</name></author><author><name sortKey="Madden, T J" uniqKey="Madden T">T.J. Madden</name></author><author><name sortKey="Miller, K L" uniqKey="Miller K">K.L. Miller</name></author><author><name sortKey="Nahrstedt, K" uniqKey="Nahrstedt K">K. Nahrstedt</name></author><author><name sortKey="Pulokas, J" uniqKey="Pulokas J">J. Pulokas</name></author><author><name sortKey="Reilein, A" uniqKey="Reilein A">A. Reilein</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Potterton, E" uniqKey="Potterton E">E. Potterton</name></author><author><name sortKey="Briggs, P" uniqKey="Briggs P">P. Briggs</name></author><author><name sortKey="Turkenburg, M" uniqKey="Turkenburg M">M. Turkenburg</name></author><author><name sortKey="Dodson, E" uniqKey="Dodson E">E. Dodson</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Punjani, A" uniqKey="Punjani A">A. Punjani</name></author><author><name sortKey="Rubinstein, J L" uniqKey="Rubinstein J">J.L. Rubinstein</name></author><author><name sortKey="Fleet, D J" uniqKey="Fleet D">D.J. Fleet</name></author><author><name sortKey="Brubaker, M A" uniqKey="Brubaker M">M.A. Brubaker</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Raj, V S" uniqKey="Raj V">V.S. Raj</name></author><author><name sortKey="Mou, H" uniqKey="Mou H">H. Mou</name></author><author><name sortKey="Smits, S L" uniqKey="Smits S">S.L. Smits</name></author><author><name sortKey="Dekkers, D H" uniqKey="Dekkers D">D.H. Dekkers</name></author><author><name sortKey="Muller, M A" uniqKey="Muller M">M.A. Müller</name></author><author><name sortKey="Dijkman, R" uniqKey="Dijkman R">R. Dijkman</name></author><author><name sortKey="Muth, D" uniqKey="Muth D">D. Muth</name></author><author><name sortKey="Demmers, J A" uniqKey="Demmers J">J.A. Demmers</name></author><author><name sortKey="Zaki, A" uniqKey="Zaki A">A. Zaki</name></author><author><name sortKey="Fouchier, R A" uniqKey="Fouchier R">R.A. Fouchier</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Rockx, B" uniqKey="Rockx B">B. Rockx</name></author><author><name sortKey="Corti, D" uniqKey="Corti D">D. Corti</name></author><author><name sortKey="Donaldson, E" uniqKey="Donaldson E">E. Donaldson</name></author><author><name sortKey="Sheahan, T" uniqKey="Sheahan T">T. Sheahan</name></author><author><name sortKey="Stadler, K" uniqKey="Stadler K">K. Stadler</name></author><author><name sortKey="Lanzavecchia, A" uniqKey="Lanzavecchia A">A. Lanzavecchia</name></author><author><name sortKey="Baric, R" uniqKey="Baric R">R. Baric</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Rosen, O" uniqKey="Rosen O">O. Rosen</name></author><author><name sortKey="Chan, L L" uniqKey="Chan L">L.L. Chan</name></author><author><name sortKey="Abiona, O M" uniqKey="Abiona O">O.M. Abiona</name></author><author><name sortKey="Gough, P" uniqKey="Gough P">P. Gough</name></author><author><name sortKey="Wang, L" uniqKey="Wang L">L. Wang</name></author><author><name sortKey="Shi, W" uniqKey="Shi W">W. Shi</name></author><author><name sortKey="Zhang, Y" uniqKey="Zhang Y">Y. Zhang</name></author><author><name sortKey="Wang, N" uniqKey="Wang N">N. Wang</name></author><author><name sortKey="Kong, W P" uniqKey="Kong W">W.P. Kong</name></author><author><name sortKey="Mclellan, J S" uniqKey="Mclellan J">J.S. McLellan</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Schultze, B" uniqKey="Schultze B">B. Schultze</name></author><author><name sortKey="Gross, H J" uniqKey="Gross H">H.J. Gross</name></author><author><name sortKey="Brossmer, R" uniqKey="Brossmer R">R. Brossmer</name></author><author><name sortKey="Herrler, G" uniqKey="Herrler G">G. Herrler</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Scobey, T" uniqKey="Scobey T">T. Scobey</name></author><author><name sortKey="Yount, B L" uniqKey="Yount B">B.L. Yount</name></author><author><name sortKey="Sims, A C" uniqKey="Sims A">A.C. Sims</name></author><author><name sortKey="Donaldson, E F" uniqKey="Donaldson E">E.F. Donaldson</name></author><author><name sortKey="Agnihothram, S S" uniqKey="Agnihothram S">S.S. Agnihothram</name></author><author><name sortKey="Menachery, V D" uniqKey="Menachery V">V.D. Menachery</name></author><author><name sortKey="Graham, R L" uniqKey="Graham R">R.L. Graham</name></author><author><name sortKey="Swanstrom, J" uniqKey="Swanstrom J">J. Swanstrom</name></author><author><name sortKey="Bove, P F" uniqKey="Bove P">P.F. Bove</name></author><author><name sortKey="Kim, J D" uniqKey="Kim J">J.D. Kim</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Shang, J" uniqKey="Shang J">J. Shang</name></author><author><name sortKey="Zheng, Y" uniqKey="Zheng Y">Y. Zheng</name></author><author><name sortKey="Yang, Y" uniqKey="Yang Y">Y. Yang</name></author><author><name sortKey="Liu, C" uniqKey="Liu C">C. Liu</name></author><author><name sortKey="Geng, Q" uniqKey="Geng Q">Q. Geng</name></author><author><name sortKey="Luo, C" uniqKey="Luo C">C. Luo</name></author><author><name sortKey="Zhang, W" uniqKey="Zhang W">W. Zhang</name></author><author><name sortKey="Li, F" uniqKey="Li F">F. Li</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Shang, J" uniqKey="Shang J">J. Shang</name></author><author><name sortKey="Zheng, Y" uniqKey="Zheng Y">Y. Zheng</name></author><author><name sortKey="Yang, Y" uniqKey="Yang Y">Y. Yang</name></author><author><name sortKey="Liu, C" uniqKey="Liu C">C. Liu</name></author><author><name sortKey="Geng, Q" uniqKey="Geng Q">Q. Geng</name></author><author><name sortKey="Tai, W" uniqKey="Tai W">W. Tai</name></author><author><name sortKey="Du, L" uniqKey="Du L">L. Du</name></author><author><name sortKey="Zhou, Y" uniqKey="Zhou Y">Y. Zhou</name></author><author><name sortKey="Zhang, W" uniqKey="Zhang W">W. Zhang</name></author><author><name sortKey="Li, F" uniqKey="Li F">F. Li</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Song, W" uniqKey="Song W">W. Song</name></author><author><name sortKey="Gui, M" uniqKey="Gui M">M. Gui</name></author><author><name sortKey="Wang, X" uniqKey="Wang X">X. Wang</name></author><author><name sortKey="Xiang, Y" uniqKey="Xiang Y">Y. Xiang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Stewart, S A" uniqKey="Stewart S">S.A. Stewart</name></author><author><name sortKey="Dykxhoorn, D M" uniqKey="Dykxhoorn D">D.M. Dykxhoorn</name></author><author><name sortKey="Palliser, D" uniqKey="Palliser D">D. Palliser</name></author><author><name sortKey="Mizuno, H" uniqKey="Mizuno H">H. Mizuno</name></author><author><name sortKey="Yu, E Y" uniqKey="Yu E">E.Y. Yu</name></author><author><name sortKey="An, D S" uniqKey="An D">D.S. An</name></author><author><name sortKey="Sabatini, D M" uniqKey="Sabatini D">D.M. Sabatini</name></author><author><name sortKey="Chen, I S" uniqKey="Chen I">I.S. Chen</name></author><author><name sortKey="Hahn, W C" uniqKey="Hahn W">W.C. Hahn</name></author><author><name sortKey="Sharp, P A" uniqKey="Sharp P">P.A. Sharp</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Suloway, C" uniqKey="Suloway C">C. Suloway</name></author><author><name sortKey="Pulokas, J" uniqKey="Pulokas J">J. Pulokas</name></author><author><name sortKey="Fellmann, D" uniqKey="Fellmann D">D. Fellmann</name></author><author><name sortKey="Cheng, A" uniqKey="Cheng A">A. Cheng</name></author><author><name sortKey="Guerra, F" uniqKey="Guerra F">F. Guerra</name></author><author><name sortKey="Quispe, J" uniqKey="Quispe J">J. Quispe</name></author><author><name sortKey="Stagg, S" uniqKey="Stagg S">S. Stagg</name></author><author><name sortKey="Potter, C S" uniqKey="Potter C">C.S. Potter</name></author><author><name sortKey="Carragher, B" uniqKey="Carragher B">B. Carragher</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Tang, G" uniqKey="Tang G">G. Tang</name></author><author><name sortKey="Peng, L" uniqKey="Peng L">L. Peng</name></author><author><name sortKey="Baldwin, P R" uniqKey="Baldwin P">P.R. Baldwin</name></author><author><name sortKey="Mann, D S" uniqKey="Mann D">D.S. Mann</name></author><author><name sortKey="Jiang, W" uniqKey="Jiang W">W. Jiang</name></author><author><name sortKey="Rees, I" uniqKey="Rees I">I. Rees</name></author><author><name sortKey="Ludtke, S J" uniqKey="Ludtke S">S.J. Ludtke</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Tang, X C" uniqKey="Tang X">X.C. Tang</name></author><author><name sortKey="Agnihothram, S S" uniqKey="Agnihothram S">S.S. Agnihothram</name></author><author><name sortKey="Jiao, Y" uniqKey="Jiao Y">Y. Jiao</name></author><author><name sortKey="Stanhope, J" uniqKey="Stanhope J">J. Stanhope</name></author><author><name sortKey="Graham, R L" uniqKey="Graham R">R.L. Graham</name></author><author><name sortKey="Peterson, E C" uniqKey="Peterson E">E.C. Peterson</name></author><author><name sortKey="Avnir, Y" uniqKey="Avnir Y">Y. Avnir</name></author><author><name sortKey="Tallarico, A S" uniqKey="Tallarico A">A.S. Tallarico</name></author><author><name sortKey="Sheehan, J" uniqKey="Sheehan J">J. Sheehan</name></author><author><name sortKey="Zhu, Q" uniqKey="Zhu Q">Q. Zhu</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Van Boheemen, S" uniqKey="Van Boheemen S">S. van Boheemen</name></author><author><name sortKey="De Graaf, M" uniqKey="De Graaf M">M. de Graaf</name></author><author><name sortKey="Lauber, C" uniqKey="Lauber C">C. Lauber</name></author><author><name sortKey="Bestebroer, T M" uniqKey="Bestebroer T">T.M. Bestebroer</name></author><author><name sortKey="Raj, V S" uniqKey="Raj V">V.S. Raj</name></author><author><name sortKey="Zaki, A M" uniqKey="Zaki A">A.M. Zaki</name></author><author><name sortKey="Osterhaus, A D" uniqKey="Osterhaus A">A.D. Osterhaus</name></author><author><name sortKey="Haagmans, B L" uniqKey="Haagmans B">B.L. Haagmans</name></author><author><name sortKey="Gorbalenya, A E" uniqKey="Gorbalenya A">A.E. Gorbalenya</name></author><author><name sortKey="Snijder, E J" uniqKey="Snijder E">E.J. Snijder</name></author><author><name sortKey="Fouchier, R A" uniqKey="Fouchier R">R.A. Fouchier</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Voss, N R" uniqKey="Voss N">N.R. Voss</name></author><author><name sortKey="Yoshioka, C K" uniqKey="Yoshioka C">C.K. Yoshioka</name></author><author><name sortKey="Radermacher, M" uniqKey="Radermacher M">M. Radermacher</name></author><author><name sortKey="Potter, C S" uniqKey="Potter C">C.S. Potter</name></author><author><name sortKey="Carragher, B" uniqKey="Carragher B">B. Carragher</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Walls, A C" uniqKey="Walls A">A.C. Walls</name></author><author><name sortKey="Tortorici, M A" uniqKey="Tortorici M">M.A. Tortorici</name></author><author><name sortKey="Bosch, B J" uniqKey="Bosch B">B.J. Bosch</name></author><author><name sortKey="Frenz, B" uniqKey="Frenz B">B. Frenz</name></author><author><name sortKey="Rottier, P J M" uniqKey="Rottier P">P.J.M. Rottier</name></author><author><name sortKey="Dimaio, F" uniqKey="Dimaio F">F. DiMaio</name></author><author><name sortKey="Rey, F A" uniqKey="Rey F">F.A. Rey</name></author><author><name sortKey="Veesler, D" uniqKey="Veesler D">D. Veesler</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Walls, A C" uniqKey="Walls A">A.C. Walls</name></author><author><name sortKey="Tortorici, M A" uniqKey="Tortorici M">M.A. Tortorici</name></author><author><name sortKey="Frenz, B" uniqKey="Frenz B">B. Frenz</name></author><author><name sortKey="Snijder, J" uniqKey="Snijder J">J. Snijder</name></author><author><name sortKey="Li, W" uniqKey="Li W">W. Li</name></author><author><name sortKey="Rey, F A" uniqKey="Rey F">F.A. Rey</name></author><author><name sortKey="Dimaio, F" uniqKey="Dimaio F">F. DiMaio</name></author><author><name sortKey="Bosch, B J" uniqKey="Bosch B">B.J. Bosch</name></author><author><name sortKey="Veesler, D" uniqKey="Veesler D">D. Veesler</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Walls, A C" uniqKey="Walls A">A.C. Walls</name></author><author><name sortKey="Xiong, X" uniqKey="Xiong X">X. Xiong</name></author><author><name sortKey="Park, Y J" uniqKey="Park Y">Y.J. Park</name></author><author><name sortKey="Tortorici, M A" uniqKey="Tortorici M">M.A. Tortorici</name></author><author><name sortKey="Snijder, J" uniqKey="Snijder J">J. Snijder</name></author><author><name sortKey="Quispe, J" uniqKey="Quispe J">J. Quispe</name></author><author><name sortKey="Cameroni, E" uniqKey="Cameroni E">E. Cameroni</name></author><author><name sortKey="Gopal, R" uniqKey="Gopal R">R. Gopal</name></author><author><name sortKey="Dai, M" uniqKey="Dai M">M. Dai</name></author><author><name sortKey="Lanzavecchia, A" uniqKey="Lanzavecchia A">A. Lanzavecchia</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Wang, L" uniqKey="Wang L">L. Wang</name></author><author><name sortKey="Shi, W" uniqKey="Shi W">W. Shi</name></author><author><name sortKey="Joyce, M G" uniqKey="Joyce M">M.G. Joyce</name></author><author><name sortKey="Modjarrad, K" uniqKey="Modjarrad K">K. Modjarrad</name></author><author><name sortKey="Zhang, Y" uniqKey="Zhang Y">Y. Zhang</name></author><author><name sortKey="Leung, K" uniqKey="Leung K">K. Leung</name></author><author><name sortKey="Lees, C R" uniqKey="Lees C">C.R. Lees</name></author><author><name sortKey="Zhou, T" uniqKey="Zhou T">T. Zhou</name></author><author><name sortKey="Yassine, H M" uniqKey="Yassine H">H.M. Yassine</name></author><author><name sortKey="Kanekiyo, M" uniqKey="Kanekiyo M">M. Kanekiyo</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Wang, L" uniqKey="Wang L">L. Wang</name></author><author><name sortKey="Shi, W" uniqKey="Shi W">W. Shi</name></author><author><name sortKey="Chappell, J D" uniqKey="Chappell J">J.D. Chappell</name></author><author><name sortKey="Joyce, M G" uniqKey="Joyce M">M.G. Joyce</name></author><author><name sortKey="Zhang, Y" uniqKey="Zhang Y">Y. Zhang</name></author><author><name sortKey="Kanekiyo, M" uniqKey="Kanekiyo M">M. Kanekiyo</name></author><author><name sortKey="Becker, M M" uniqKey="Becker M">M.M. Becker</name></author><author><name sortKey="Van Doremalen, N" uniqKey="Van Doremalen N">N. van Doremalen</name></author><author><name sortKey="Fischer, R" uniqKey="Fischer R">R. Fischer</name></author><author><name sortKey="Wang, N" uniqKey="Wang N">N. Wang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Wang, N" uniqKey="Wang N">N. Wang</name></author><author><name sortKey="Shi, X" uniqKey="Shi X">X. Shi</name></author><author><name sortKey="Jiang, L" uniqKey="Jiang L">L. Jiang</name></author><author><name sortKey="Zhang, S" uniqKey="Zhang S">S. Zhang</name></author><author><name sortKey="Wang, D" uniqKey="Wang D">D. Wang</name></author><author><name sortKey="Tong, P" uniqKey="Tong P">P. Tong</name></author><author><name sortKey="Guo, D" uniqKey="Guo D">D. Guo</name></author><author><name sortKey="Fu, L" uniqKey="Fu L">L. Fu</name></author><author><name sortKey="Cui, Y" uniqKey="Cui Y">Y. Cui</name></author><author><name sortKey="Liu, X" uniqKey="Liu X">X. Liu</name></author></analytic></biblStruct><biblStruct></biblStruct><biblStruct><analytic><author><name sortKey="Xiong, X" uniqKey="Xiong X">X. Xiong</name></author><author><name sortKey="Tortorici, M A" uniqKey="Tortorici M">M.A. Tortorici</name></author><author><name sortKey="Snijder, J" uniqKey="Snijder J">J. Snijder</name></author><author><name sortKey="Yoshioka, C" uniqKey="Yoshioka C">C. Yoshioka</name></author><author><name sortKey="Walls, A C" uniqKey="Walls A">A.C. Walls</name></author><author><name sortKey="Li, W" uniqKey="Li W">W. Li</name></author><author><name sortKey="Mcguire, A T" uniqKey="Mcguire A">A.T. McGuire</name></author><author><name sortKey="Rey, F A" uniqKey="Rey F">F.A. Rey</name></author><author><name sortKey="Bosch, B J" uniqKey="Bosch B">B.J. Bosch</name></author><author><name sortKey="Veesler, D" uniqKey="Veesler D">D. Veesler</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Ying, T" uniqKey="Ying T">T. Ying</name></author><author><name sortKey="Du, L" uniqKey="Du L">L. Du</name></author><author><name sortKey="Ju, T W" uniqKey="Ju T">T.W. Ju</name></author><author><name sortKey="Prabakaran, P" uniqKey="Prabakaran P">P. Prabakaran</name></author><author><name sortKey="Lau, C C" uniqKey="Lau C">C.C. Lau</name></author><author><name sortKey="Lu, L" uniqKey="Lu L">L. Lu</name></author><author><name sortKey="Liu, Q" uniqKey="Liu Q">Q. Liu</name></author><author><name sortKey="Wang, L" uniqKey="Wang L">L. Wang</name></author><author><name sortKey="Feng, Y" uniqKey="Feng Y">Y. Feng</name></author><author><name sortKey="Wang, Y" uniqKey="Wang Y">Y. Wang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Yuan, Y" uniqKey="Yuan Y">Y. Yuan</name></author><author><name sortKey="Cao, D" uniqKey="Cao D">D. Cao</name></author><author><name sortKey="Zhang, Y" uniqKey="Zhang Y">Y. Zhang</name></author><author><name sortKey="Ma, J" uniqKey="Ma J">J. Ma</name></author><author><name sortKey="Qi, J" uniqKey="Qi J">J. Qi</name></author><author><name sortKey="Wang, Q" uniqKey="Wang Q">Q. Wang</name></author><author><name sortKey="Lu, G" uniqKey="Lu G">G. Lu</name></author><author><name sortKey="Wu, Y" uniqKey="Wu Y">Y. Wu</name></author><author><name sortKey="Yan, J" uniqKey="Yan J">J. Yan</name></author><author><name sortKey="Shi, Y" uniqKey="Shi Y">Y. Shi</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Yusof, M F" uniqKey="Yusof M">M.F. Yusof</name></author><author><name sortKey="Queen, K" uniqKey="Queen K">K. Queen</name></author><author><name sortKey="Eltahir, Y M" uniqKey="Eltahir Y">Y.M. Eltahir</name></author><author><name sortKey="Paden, C R" uniqKey="Paden C">C.R. Paden</name></author><author><name sortKey="Al Hammadi, Z M A H" uniqKey="Al Hammadi Z">Z.M.A.H. Al Hammadi</name></author><author><name sortKey="Tao, Y" uniqKey="Tao Y">Y. Tao</name></author><author><name sortKey="Li, Y" uniqKey="Li Y">Y. Li</name></author><author><name sortKey="Khalafalla, A I" uniqKey="Khalafalla A">A.I. Khalafalla</name></author><author><name sortKey="Shi, M" uniqKey="Shi M">M. Shi</name></author><author><name sortKey="Zhang, J" uniqKey="Zhang J">J. Zhang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Zaki, A M" uniqKey="Zaki A">A.M. Zaki</name></author><author><name sortKey="Van Boheemen, S" uniqKey="Van Boheemen S">S. van Boheemen</name></author><author><name sortKey="Bestebroer, T M" uniqKey="Bestebroer T">T.M. Bestebroer</name></author><author><name sortKey="Osterhaus, A D" uniqKey="Osterhaus A">A.D. Osterhaus</name></author><author><name sortKey="Fouchier, R A" uniqKey="Fouchier R">R.A. Fouchier</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Zhang, K" uniqKey="Zhang K">K. Zhang</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Zheng, S Q" uniqKey="Zheng S">S.Q. Zheng</name></author><author><name sortKey="Palovcak, E" uniqKey="Palovcak E">E. Palovcak</name></author><author><name sortKey="Armache, J P" uniqKey="Armache J">J.P. Armache</name></author><author><name sortKey="Verba, K A" uniqKey="Verba K">K.A. Verba</name></author><author><name sortKey="Cheng, Y" uniqKey="Cheng Y">Y. Cheng</name></author><author><name sortKey="Agard, D A" uniqKey="Agard D">D.A. Agard</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Zivanov, J" uniqKey="Zivanov J">J. Zivanov</name></author><author><name sortKey="Nakane, T" uniqKey="Nakane T">T. Nakane</name></author><author><name sortKey="Forsberg, B O" uniqKey="Forsberg B">B.O. Forsberg</name></author><author><name sortKey="Kimanius, D" uniqKey="Kimanius D">D. Kimanius</name></author><author><name sortKey="Hagen, W J" uniqKey="Hagen W">W.J. Hagen</name></author><author><name sortKey="Lindahl, E" uniqKey="Lindahl E">E. Lindahl</name></author><author><name sortKey="Scheres, S H" uniqKey="Scheres S">S.H. Scheres</name></author></analytic></biblStruct></listBibl></div1></back></TEI><pmc article-type="research-article"><pmc-dir>properties open_access</pmc-dir>
<front><journal-meta><journal-id journal-id-type="nlm-ta">Cell Rep</journal-id><journal-id journal-id-type="iso-abbrev">Cell Rep</journal-id><journal-title-group><journal-title>Cell Reports</journal-title></journal-title-group><issn pub-type="epub">2211-1247</issn><publisher><publisher-name>Cell Press</publisher-name></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">31553909</article-id><article-id pub-id-type="pmc">6935267</article-id><article-id pub-id-type="publisher-id">S2211-1247(19)31100-3</article-id><article-id pub-id-type="doi">10.1016/j.celrep.2019.08.052</article-id><article-categories><subj-group subj-group-type="heading"><subject>Article</subject></subj-group></article-categories><title-group><article-title>Structural Definition of a Neutralization-Sensitive Epitope on the MERS-CoV S1-NTD</article-title></title-group><contrib-group><contrib contrib-type="author" id="au1"><name><surname>Wang</surname><given-names>Nianshuang</given-names></name><xref rid="aff1" ref-type="aff">1</xref></contrib><contrib contrib-type="author" id="au2"><name><surname>Rosen</surname><given-names>Osnat</given-names></name><xref rid="aff2" ref-type="aff">2</xref><xref rid="fn1" ref-type="fn">6</xref></contrib><contrib contrib-type="author" id="au3"><name><surname>Wang</surname><given-names>Lingshu</given-names></name><xref rid="aff2" ref-type="aff">2</xref></contrib><contrib contrib-type="author" id="au4"><name><surname>Turner</surname><given-names>Hannah L.</given-names></name><xref rid="aff3" ref-type="aff">3</xref></contrib><contrib contrib-type="author" id="au5"><name><surname>Stevens</surname><given-names>Laura J.</given-names></name><xref rid="aff4" ref-type="aff">4</xref></contrib><contrib contrib-type="author" id="au6"><name><surname>Corbett</surname><given-names>Kizzmekia S.</given-names></name><xref rid="aff2" ref-type="aff">2</xref></contrib><contrib contrib-type="author" id="au7"><name><surname>Bowman</surname><given-names>Charles A.</given-names></name><xref rid="aff3" ref-type="aff">3</xref></contrib><contrib contrib-type="author" id="au8"><name><surname>Pallesen</surname><given-names>Jesper</given-names></name><xref rid="aff3" ref-type="aff">3</xref></contrib><contrib contrib-type="author" id="au9"><name><surname>Shi</surname><given-names>Wei</given-names></name><xref rid="aff2" ref-type="aff">2</xref></contrib><contrib contrib-type="author" id="au10"><name><surname>Zhang</surname><given-names>Yi</given-names></name><xref rid="aff2" ref-type="aff">2</xref></contrib><contrib contrib-type="author" id="au11"><name><surname>Leung</surname><given-names>Kwanyee</given-names></name><xref rid="aff2" ref-type="aff">2</xref></contrib><contrib contrib-type="author" id="au12"><name><surname>Kirchdoerfer</surname><given-names>Robert N.</given-names></name><xref rid="aff3" ref-type="aff">3</xref></contrib><contrib contrib-type="author" id="au13"><name><surname>Becker</surname><given-names>Michelle M.</given-names></name><xref rid="aff4" ref-type="aff">4</xref></contrib><contrib contrib-type="author" id="au14"><name><surname>Denison</surname><given-names>Mark R.</given-names></name><xref rid="aff4" ref-type="aff">4</xref><xref rid="aff5" ref-type="aff">5</xref></contrib><contrib contrib-type="author" id="au15"><name><surname>Chappell</surname><given-names>James D.</given-names></name><xref rid="aff4" ref-type="aff">4</xref></contrib><contrib contrib-type="author" id="au16"><name><surname>Ward</surname><given-names>Andrew B.</given-names></name><xref rid="aff3" ref-type="aff">3</xref></contrib><contrib contrib-type="author" id="au17"><name><surname>Graham</surname><given-names>Barney S.</given-names></name><xref rid="aff2" ref-type="aff">2</xref></contrib><contrib contrib-type="author" id="au18"><name><surname>McLellan</surname><given-names>Jason S.</given-names></name><email>jmclellan@austin.utexas.edu</email><xref rid="aff1" ref-type="aff">1</xref><xref rid="fn2" ref-type="fn">7</xref><xref rid="cor1" ref-type="corresp">∗</xref></contrib></contrib-group><aff id="aff1"><label>1</label>Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX 78712, USA</aff><aff id="aff2"><label>2</label>Vaccine Research Center, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892, USA</aff><aff id="aff3"><label>3</label>Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA</aff><aff id="aff4"><label>4</label>Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</aff><aff id="aff5"><label>5</label>Department of Pathology, Microbiology, and Immunology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA</aff><author-notes><corresp id="cor1"><label>∗</label>Corresponding author <email>jmclellan@austin.utexas.edu</email></corresp><fn id="fn1"><label>6</label><p id="ntpara0010">Present address: Department of Biotechnology, Israel Institute for Biological Research, Ness-ziona, Israel</p></fn><fn id="fn2"><label>7</label><p id="ntpara0015">Lead Contact</p></fn></author-notes><pub-date pub-type="pmc-release"><day>24</day><month>9</month><year>2019</year></pub-date><pmc-comment> PMC Release delay is 0 months and 0 days and was based on .</pmc-comment>
<pub-date pub-type="ppub"><day>24</day><month>9</month><year>2019</year></pub-date><pub-date pub-type="epub"><day>24</day><month>9</month><year>2019</year></pub-date><volume>28</volume><issue>13</issue><fpage>3395</fpage><lpage>3405.e6</lpage><history><date date-type="received"><day>6</day><month>2</month><year>2019</year></date><date date-type="rev-recd"><day>6</day><month>6</month><year>2019</year></date><date date-type="accepted"><day>16</day><month>8</month><year>2019</year></date></history><permissions><copyright-statement>© 2019 The Author(s)</copyright-statement><copyright-year>2019</copyright-year><license><license-p>Since January 2020 Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre - including this research content - immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.</license-p></license></permissions><abstract id="abs0010"><title>Summary</title><p>Middle East respiratory syndrome coronavirus (MERS-CoV) emerged into the human population in 2012 and has caused substantial morbidity and mortality. Potently neutralizing antibodies targeting the receptor-binding domain (RBD) on MERS-CoV spike (S) protein have been characterized, but much less is known about antibodies targeting non-RBD epitopes. Here, we report the structural and functional characterization of G2, a neutralizing antibody targeting the MERS-CoV S1 N-terminal domain (S1-NTD). Structures of G2 alone and in complex with the MERS-CoV S1-NTD define a site of vulnerability comprising two loops, each of which contain a residue mutated in G2-escape variants. Cell-surface binding studies and <italic>in vitro</italic> competition experiments demonstrate that G2 strongly disrupts the attachment of MERS-CoV S to its receptor, dipeptidyl peptidase-4 (DPP4), with the inhibition requiring the native trimeric S conformation. These results advance our understanding of antibody-mediated neutralization of coronaviruses and should facilitate the development of immunotherapeutics and vaccines against MERS-CoV.</p></abstract><abstract abstract-type="graphical" id="abs0015"><title>Graphical Abstract</title><fig id="undfig1" position="anchor"><graphic xlink:href="fx1_lrg"></graphic></fig></abstract><abstract abstract-type="author-highlights" id="abs0020"><title>Highlights</title><p><list list-type="simple" id="ulist0010"><list-item id="u0010"><label>•</label><p id="p0010">The epitope for the neutralizing antibody G2 is confined to the apex of the MERS-CoV S1-NTD</p></list-item><list-item id="u0015"><label>•</label><p id="p0015">G2 epitope is relatively well conserved</p></list-item><list-item id="u0020"><label>•</label><p id="p0020">G2 IgG and Fab both neutralize pseudotyped and authentic MERS-CoV</p></list-item><list-item id="u0025"><label>•</label><p id="p0025">G2 neutralizes by preventing the binding of DPP4 to trimeric S protein</p></list-item></list></p></abstract><abstract abstract-type="teaser" id="abs0025"><p>Wang et al. report the structural and functional characterization of the Middle East respiratory syndrome coronavirus (MERS-CoV)-neutralizing antibody G2. G2 recognizes a conserved epitope on the MERS-CoV S1 N-terminal domain (S1-NTD) and neutralizes MERS-CoV by interfering with binding to host receptor dipeptidyl peptidase-4 (DPP4). The findings are relevant for understanding the viral attachment mechanism and for the development of S1-NTD-based vaccines.</p></abstract><kwd-group id="kwrds0010"><title>Keywords</title><kwd>MERS-CoV</kwd><kwd>coronavirus</kwd><kwd>crystal structure</kwd><kwd>electron microscopy</kwd><kwd>DPP4</kwd><kwd>receptor-binding</kwd><kwd>membrane fusion</kwd></kwd-group></article-meta><notes><p id="misc0010">Published: September 24, 2019</p></notes></front><body><sec id="sec1"><title>Introduction</title><p id="p0030">Middle East respiratory syndrome coronavirus (MERS-CoV) is a zoonotic coronavirus first identified in Saudi Arabia in 2012 (<xref rid="bib63" ref-type="bibr">van Boheemen et al., 2012</xref>, <xref rid="bib76" ref-type="bibr">Zaki et al., 2012</xref>). MERS-CoV can cause severe acute respiratory disease in humans with symptoms including fever, cough, and shortness of breath (<xref rid="bib71" ref-type="bibr">WHO, 2018</xref>). Through the end of 2018, the World Health Organization (WHO) has been notified of 2,266 laboratory-confirmed cases of MERS-CoV infection from 27 countries, with most cases occurring in the Middle East (<xref rid="bib71" ref-type="bibr">WHO, 2018</xref>). The MERS-CoV case-fatality rate for laboratory-confirmed severe disease is 36%, with the number of deaths exceeding 800. MERS-CoV likely originated from bats, with camels functioning as a secondary or intermediate host (<xref rid="bib4" ref-type="bibr">Azhar et al., 2014</xref>, <xref rid="bib39" ref-type="bibr">Mohd et al., 2016</xref>). Small clusters of infections in several countries suggested that limited human-to-human transmission can occur through close contact (<xref rid="bib23" ref-type="bibr">Ki, 2015</xref>, <xref rid="bib42" ref-type="bibr">Oboho et al., 2015</xref>). Due to the ongoing circulation, high pathogenicity, and capacity for inter-human transmission associated with MERS-CoV, there is a persistent concern about a possible pandemic. Because no specific antiviral drugs or protective vaccines are currently available, efficient countermeasures against this virus are urgently needed.</p><p id="p0035">The surface of coronavirus virions is decorated with the large trimeric spike (S) glycoprotein, which mediates cell entry (<xref rid="bib17" ref-type="bibr">Gierer et al., 2013</xref>, <xref rid="bib30" ref-type="bibr">Li, 2016</xref>). The MERS-CoV S glycoprotein is synthesized as a single-chain precursor that is subsequently cleaved by furin-like host proteases to generate the S1 and S2 subunits (<xref rid="bib37" ref-type="bibr">Millet and Whittaker, 2014</xref>). The mature S protein is a homotrimer of non-covalently associated S1 and S2 subunits whereby a trimer of S1 acts as a fusion-suppressive cap and sits atop a trimer of S2 subunits. Binding of S1 to the host receptor dipeptidyl peptidase-4 (DPP4) (<xref rid="bib35" ref-type="bibr">Lu et al., 2013</xref>, <xref rid="bib51" ref-type="bibr">Raj et al., 2013</xref>, <xref rid="bib70" ref-type="bibr">Wang et al., 2013</xref>) initiates a large irreversible conformational change of S2, which mediates fusion of the viral and host-cell membranes. Cryoelectron microscopy (cryo-EM) structures of various β-coronaviruses (<xref rid="bib19" ref-type="bibr">Gui et al., 2017</xref>, <xref rid="bib24" ref-type="bibr">Kirchdoerfer et al., 2016</xref>, <xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>, <xref rid="bib56" ref-type="bibr">Shang et al., 2018a</xref>, <xref rid="bib57" ref-type="bibr">Shang et al., 2018b</xref>, <xref rid="bib65" ref-type="bibr">Walls et al., 2016a</xref>, <xref rid="bib66" ref-type="bibr">Walls et al., 2016b</xref>, <xref rid="bib72" ref-type="bibr">Xiong et al., 2018</xref>, <xref rid="bib74" ref-type="bibr">Yuan et al., 2017</xref>) have revealed a four-domain architecture of S1 including an N-terminal domain (NTD), a C-terminal domain (CTD), and subdomains I and II. S1-NTD or S1-CTD can function as the receptor-binding domain (RBD) depending on the specific coronavirus. Most β-coronaviruses, including severe acute respiratory syndrome-CoV (SARS-CoV) (<xref rid="bib32" ref-type="bibr">Li et al., 2003</xref>, <xref rid="bib31" ref-type="bibr">Li et al., 2005</xref>) and MERS-CoV (<xref rid="bib35" ref-type="bibr">Lu et al., 2013</xref>, <xref rid="bib51" ref-type="bibr">Raj et al., 2013</xref>, <xref rid="bib70" ref-type="bibr">Wang et al., 2013</xref>), use the S1-CTD to bind to their functional receptor, whereas some lineage A β-coronaviruses, such as mouse hepatitis virus (MHV) (<xref rid="bib45" ref-type="bibr">Peng et al., 2011</xref>) and bovine coronavirus (BCoV) (<xref rid="bib46" ref-type="bibr">Peng et al., 2012</xref>), bind receptors using the S1-NTD.</p><p id="p0040">Prefusion S1 proteins from some coronaviruses, including human coronavirus HKU1 (HCoV-HKU1) (<xref rid="bib24" ref-type="bibr">Kirchdoerfer et al., 2016</xref>) and MHV (<xref rid="bib65" ref-type="bibr">Walls et al., 2016a</xref>), fold into a well-packed symmetric trimer with NTDs and CTDs tightly interacting with each other. In this conformation, the receptor-binding surface on the CTDs is mostly occluded within the internal surface of the trimer. Conversely, prefusion SARS-CoV (<xref rid="bib19" ref-type="bibr">Gui et al., 2017</xref>, <xref rid="bib74" ref-type="bibr">Yuan et al., 2017</xref>) and MERS-CoV (<xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>, <xref rid="bib74" ref-type="bibr">Yuan et al., 2017</xref>) S1 proteins adopt dynamic open and closed conformations, wherein each of the three S1-CTDs adopt either a compact “down” conformation that buries the receptor-binding surface, or an “up” conformation that facilitates binding with host-cell receptors. It has been hypothesized that these conformations exist in an equilibrium, with receptor binding to the up conformation resulting in a three CTD up arrangement that is unstable, resulting in dissociation of S1 and refolding of S2 (<xref rid="bib19" ref-type="bibr">Gui et al., 2017</xref>, <xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>, <xref rid="bib58" ref-type="bibr">Song et al., 2018</xref>, <xref rid="bib67" ref-type="bibr">Walls et al., 2019</xref>, <xref rid="bib74" ref-type="bibr">Yuan et al., 2017</xref>).</p><p id="p0045">Coronavirus S1-NTDs adopt a three-layer structure, with a core region formed by a galectin-like β sandwich fold, a top region above the core that is commonly used by some lineage A β-coronaviruses to bind proteins or glycan receptors, and a bottom conserved region that stretches out to connect with the S1-CTD (<xref rid="bib45" ref-type="bibr">Peng et al., 2011</xref>). Structures of MERS-CoV S1-NTD have been reported previously (<xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>, <xref rid="bib74" ref-type="bibr">Yuan et al., 2017</xref>), but specific interactions with host-cell factors have not been well characterized. Recent studies have suggested that CEACAM5 (<xref rid="bib6" ref-type="bibr">Chan et al., 2016</xref>), GRP78 (<xref rid="bib8" ref-type="bibr">Chu et al., 2018</xref>), and sialic acid (<xref rid="bib33" ref-type="bibr">Li et al., 2017</xref>) may serve as important attachment factors prior to DPP4 binding, and these interactions may be mediated by the S1-NTD.</p><p id="p0050">As the main protein on the surface of the coronavirus virion, the S protein is the key target for protective antibody responses (<xref rid="bib30" ref-type="bibr">Li, 2016</xref>, <xref rid="bib38" ref-type="bibr">Modjarrad et al., 2016</xref>). Many neutralizing antibodies targeting the MERS-CoV S1-CTD, which is the RBD, have been isolated and characterized (<xref rid="bib10" ref-type="bibr">Corti et al., 2015</xref>, <xref rid="bib22" ref-type="bibr">Jiang et al., 2014</xref>, <xref rid="bib34" ref-type="bibr">Li et al., 2015</xref>, <xref rid="bib62" ref-type="bibr">Tang et al., 2014</xref>, <xref rid="bib73" ref-type="bibr">Ying et al., 2014</xref>), and structural studies have revealed that their epitopes overlap with the DPP4-binding surface, thereby providing a structural basis for neutralization. Besides the immunodominant RBD, the S1-NTD has also been shown to induce protective antibody responses in a mouse model (<xref rid="bib21" ref-type="bibr">Jiaming et al., 2017</xref>). Neutralizing antibodies targeting MERS-CoV S1-NTD have been reported, including human antibody CDC2-A2, murine antibodies G2 and 5F9, and macaque antibodies FIB-H1 and JC57-13 (<xref rid="bib7" ref-type="bibr">Chen et al., 2017</xref>, <xref rid="bib68" ref-type="bibr">Wang et al., 2015</xref>, <xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>). Of these antibodies, G2 is the most potent, with broad neutralization potential against an array of MERS-CoV strains (<xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>). Additionally, G2 and other NTD-specific MERS-CoV antibodies have been shown to confer protection against lethal challenge in animal models (<xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>). However, the lack of structural information for G2 has hindered the definition of its epitope and determination of its mechanism of action.</p><p id="p0055">To address this knowledge gap, we initiated a series of comprehensive studies. Here, we report the crystal structures of G2 Fab alone and bound to the MERS-CoV S1-NTD, as well as the results from biochemical, biophysical, and cell-based assays. These studies define a site of vulnerability on the MERS-CoV spike and elucidate a mechanism of neutralization that involves inhibition of attachment to DPP4.</p></sec><sec id="sec2"><title>Results</title><sec id="sec2.1"><title>The G2 Epitope Is Confined Solely to the S1-NTD</title><p id="p0060">We conducted surface plasmon resonance (SPR) experiments to characterize the interaction between G2 Fab and MERS-CoV S1-NTD (<xref rid="fig1" ref-type="fig">Figure 1</xref>A), as well as the interaction between G2 Fab and the prefusion-stabilized MERS-CoV S ectodomain (MERS-CoV S-2P) (<xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>) (<xref rid="fig1" ref-type="fig">Figure 1</xref>B). The affinities of G2 Fab for S1-NTD and MERS-CoV S-2P were very similar, with <italic>K</italic><sub>D</sub>s of 28.0 and 30.3 nM, respectively. The binding kinetics were also in good agreement, indicating that the G2 epitope is confined to the S1-NTD.<fig id="fig1"><label>Figure 1</label><caption><p>Two Selected G2-Escape Mutations and Their Impact on G2 Binding and Neutralization</p><p>(A–D) Binding of G2 Fab to immobilized (A) MERS-CoV S1-NTD, (B) MERS-CoV S-2P, (C) MERS-CoV S-2P-S28F, and (D) MERS-CoV S-2P-G198D measured by surface plasmon resonance (SPR). The same concentration series of G2 Fab was used in (A)–(D). Best global fit of the data to a 1:1 binding model is shown as colored lines.</p><p>(E) Neutralization activity of G2 IgG was measured against pseudotyped lentivirus bearing MERS-CoV S (WT) and two variants (S28F and G198D). Percent neutralization of WT (red), S28F (blue), and G198D (black) S pseudovirions at the different antibody concentrations is shown. Data points represent the mean of three technical replicates with standard errors.</p><p>(F) Neutralization activity of G2 IgG was measured against authentic MERS-CoV (WT) and the G198D variant. Percent neutralization of WT (red) and G198D (black) MERS-CoV at the different antibody concentrations is shown. Data points for the WT virus represent the mean of two technical replicates.</p></caption><graphic xlink:href="gr1_lrg"></graphic></fig></p><p id="p0065">To more precisely localize the G2 epitope on the S1-NTD, we performed <italic>in vitro</italic> selection for G2-escape variants by serial passage of recombinant MERS-CoV strain EMC/2012 in Vero 81 cell cultures (<xref rid="bib55" ref-type="bibr">Scobey et al., 2013</xref>) supplemented with progressively escalating concentrations of G2 immunoglobulin G (IgG). After 10 passages, 15 G2-resistant MERS-CoV isolates were plaque cloned and the mutations were analyzed. All 15 clones contained either an S28F or G198D substitution (<xref rid="mmc1" ref-type="supplementary-material">Table S1</xref>), suggesting that these two residues are crucial for G2 binding. To test this hypothesis, we generated MERS-CoV S-2P variants harboring the S28F or G198D substitutions. SPR measurements revealed that the S28F or G198D substitutions completely abolished binding to G2 Fab at the concentrations tested (<xref rid="fig1" ref-type="fig">Figures 1</xref>C and 1D). We next generated pseudotyped lentiviruses bearing the mutant MERS-CoV S glycoproteins (S28F or G198D) to assess the effect of these substitutions on the neutralizing activity of G2 IgG. As expected, G2 IgG potently neutralized pseudoviruses bearing wild-type (WT) S protein, whereas G2 IgG poorly neutralized the pseudoviruses harboring the escape mutations, with a maximum neutralized fraction below 45% (<xref rid="fig1" ref-type="fig">Figure 1</xref>E). We additionally verified the neutralizing ability of G2 IgG against the recombinant MERS-CoV strain EMC/2012 (<xref rid="bib2" ref-type="bibr">Almazán et al., 2013</xref>) and the G198D variant in a biosafety level 3 (BSL-3) setting. G2 IgG neutralized authentic MERS-CoV with an EC<sub>50</sub> = 0.12 nM, whereas the ability of G2 IgG to neutralize the G198D variant was barely detectable until we raised the concentration to 10 μM (<xref rid="fig1" ref-type="fig">Figure 1</xref>F). These results indicate that the G2 epitope is localized to a surface on the S1-NTD near residues Ser28 and Gly198.</p></sec><sec id="sec2.2"><title>The G2 Epitope Is Primarily Localized to Two Loops at the Top of S1-NTD</title><p id="p0070">To precisely define the G2 epitope, we determined the crystal structures of G2 Fab alone and in complex with MERS-CoV S1-NTD. G2 Fab formed crystals in space group <italic>P</italic>2<sub>1</sub> that diffracted X-rays to 2.1-Å resolution. Phasing by molecular replacement revealed four Fab molecules per asymmetric unit. After iterative rounds of model building and refinement, the final model had an R/R<sub>free</sub> = 18.1%/22.5% (<xref rid="mmc1" ref-type="supplementary-material">Table S2</xref>). Crystallization of G2 Fab in complex with the S1-NTD proved difficult and necessitated exploration of different expression and purification strategies. Ultimately, crystals of the complex were obtained in space group <italic>P</italic>2<sub>1</sub> that diffracted X-rays to 2.3-Å resolution. The molecular replacement solution contained two molecules of the complex per asymmetric unit, and iterative rounds of model building and refinement produced a final model with an R/R<sub>free</sub> = 18.0%/21.3%.</p><p id="p0075">The structure of the MERS-CoV S1-NTD is similar to that of other coronaviruses and can be separated into top, core, and bottom regions (<xref rid="fig2" ref-type="fig">Figure 2</xref>A). The MERS-CoV S1-NTD core contains a β sandwich structure formed by two β sheets containing a total of 11 β strands (<xref rid="fig2" ref-type="fig">Figure 2</xref>B, labeled β1-β11), providing a scaffold to support the top and bottom regions. Stretching out from β5, a loop (residues 189-202) designated loop2 spirals 540° perpendicularly to the β sheet, like a right-handed pseudo-helix, and comprises the majority of the G2 binding interface (∼384 Å<sup>2</sup>). A disulfide bond between Cys30 and Cys195 connects loop2 with an <italic>N</italic>-terminal loop (residues 18–33) designated loop1, which contributes the remaining binding interface with G2 (∼295 Å<sup>2</sup>) (<xref rid="fig2" ref-type="fig">Figures 2</xref>B and 2C).<fig id="fig2"><label>Figure 2</label><caption><p>Structure of G2 Fab Complexed with MERS-CoV S1-NTD</p><p>(A) Overall structure of the complex. G2 heavy and light chains are colored dark blue and white, respectively. S1-NTD is separated into top (red), core (orange), and bottom (yellow) regions. Residues Ser28 and Gly198 are shown as semi-transparent molecular surfaces.</p><p>(B) The structure of S1-NTD top region is presented in ribbon representation. The 11 β strands in the core region are labeled β1-β11. Disulfide bonds are drawn as yellow sticks.</p><p>(C) The structure of S1-NTD top region is presented as a molecular surface, viewed in the same orientation as in (B). Surfaces on loop1 and loop2 buried at the interface with G2 are encircled by a dotted line and residues forming hydrogen bonds with G2 are colored white and blue, respectively.</p><p>(D) G2 contacts two loops (loop1 and loop2) on the S1-NTD top region. Antibody complementarity-determining regions (CDRs) involved in the binding are labeled. Residues contributing to the interaction are shown in a stick representation. Hydrogen bonds and salt bridges are depicted as black dotted lines.</p><p>(E) Linear sequences of loop1 and loop2. S1-NTD residues that make hydrogen bonds to G2 are denoted with symbols. Two G2-escape mutations (S28P and G198D) naturally occurring in S proteins from two different MERS-CoV strains (camel/UAE_B42 and Riyadh_2014KSA_349, respectively) are colored red.</p></caption><graphic xlink:href="gr2_lrg"></graphic></fig></p><p id="p0080">Four of the six G2 complementarity-determining regions (CDRH1, CDRH2, CDRH3, and CDRL3) are involved in binding to the S1-NTD (<xref rid="fig2" ref-type="fig">Figure 2</xref>D). CDRL3 interacts with loop1, whereas the heavy-chain CDRs interact with loop2, except for one hydrogen bond formed between CDRH3 residue Ser97 and S1-NTD loop1 residue Ser28. G2 heavy-chain residues Tyr33, Thr54, and Ser97 form hydrogen bonds with main-chain atoms on loop2, whereas heavy-chain residues Trp50 and Tyr52 interact with side chains of loop1 residues Ser191 and Asn199, respectively. The interaction between the G2 light chain and loop1 is mediated by three residues on CDRL3 (Ser91, Glu92, and Glu93) and two residues on loop1 (Lys27 and Ser28). Loop1 residue Lys27 forms two salt bridges with Glu92 and Glu93 on CDRL3, whereas Ser28 forms hydrogen bonds with main-chain atoms of CDRL3 residues Ser91 and Glu92.</p><p id="p0085">There are eight <italic>N</italic>-linked glycosylation sites within the S1-NTD. As the crystallized protein was not treated with glycosidases, electron density for large glycan moieties can be observed on several glycosylation sites. However, none of these glycosylation sites are within the G2 binding interface (<xref rid="mmc1" ref-type="supplementary-material">Figure S1</xref>A). Thus, it is unlikely that G2 binding is dependent on the presence of <italic>N</italic>-linked glycans. To verify, SPR studies confirmed that deglycosylated S1-NTD retained a similar affinity to G2 as glycosylated S1-NTD (<xref rid="mmc1" ref-type="supplementary-material">Figure S1</xref>B).</p><p id="p0090">The structure is consistent with the G2-escape data, which showed that MERS-CoV cultured under G2 selection accumulates substitutions S28F and G198D at the G2 binding interface (<xref rid="mmc1" ref-type="supplementary-material">Table S1</xref>). Based on the structure, we predict that the S28F substitution would largely eliminate the hydrophilic interactions between loop1 and CDRL3. The G198D substitution would introduce a long side chain leading to a steric clash that would impair the interaction. In addition to these two escape mutations, we further probed the interface by introducing single mutations K27A or S191A into the S1-NTD. As expected from the structure, either of these two substitutions largely abolished the binding to G2 Fab (<xref rid="mmc1" ref-type="supplementary-material">Figure S2</xref>A).</p></sec><sec id="sec2.3"><title>The G2 Epitope Is Relatively Conserved</title><p id="p0095">Previous neutralization data demonstrated that G2 IgG can neutralize pseudoviruses with S proteins from eight different MERS-CoV strains with inhibitory concentration (IC)<sub>50</sub> values ranging from 0.010 to 0.028 μg/ml (<xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>). This is in contrast to other S1-NTD-specific antibodies, like A2 and JC57-13, which showed weaker neutralizing potency. We analyzed the S1-NTD sequences from all eight tested strains and identified amino acid differences at seven positions. However, none of these are involved in the interaction with G2 (<xref rid="mmc1" ref-type="supplementary-material">Figure S2</xref>B).</p><p id="p0100">We additionally analyzed all available MERS-CoV S sequences in GenBank. Most of the G2-interacting residues, including Lys27, Ser191, Asn193, Ala197, and Asn199, are conserved among all 232 sequences, explaining why G2 IgG can broadly neutralize MERS-CoV strains. Interestingly, we did find some MERS-CoV strains with substitutions at Ser28 or Gly198 (<xref rid="fig2" ref-type="fig">Figure 2</xref>E), the two residues that were substituted in escape variants under <italic>in vitro</italic> G2 selection. Eight sequences (ASU90362, ASU90142, ASU89988, ASU91208, ASU91284, ASU90186, ASU90010, and ASU89966) isolated from camels (<xref rid="bib75" ref-type="bibr">Yusof et al., 2017</xref>), along with one sequence isolated from a patient in 2015 (ALJ54461) (<xref rid="bib3" ref-type="bibr">Assiri et al., 2016</xref>), harbor a S28P substitution. One human MERS-CoV sequence isolated in 2014 (<xref rid="bib12" ref-type="bibr">Drosten et al., 2015</xref>) is the only one that harbors a G198D mutation. We tested an S1-NTD construct bearing the S28P substitution for binding to G2 Fab. The affinity was ∼10-fold lower compared to the affinity of WT S1-NTD (<xref rid="mmc1" ref-type="supplementary-material">Figure S2</xref>A). The natural occurrence of S28P and G198D may indicate that MERS-CoV is under selective pressure exerted by host G2-like antibody responses.</p></sec><sec id="sec2.4"><title>G2 Binding to the Prefusion Spike</title><p id="p0105">To further investigate G2 binding in the context of the MERS-CoV S trimer, we purified the MERS-CoV S0 ectodomain in complex with G2 Fab and performed negative-stain EM analysis. 2D classification suggested that the sample was heterogeneous, and postfusion rod-like particles were abundant (<xref rid="mmc1" ref-type="supplementary-material">Figure S3</xref>A), suggesting that G2 Fab is not able to prevent the prefusion-to-postfusion transformation of S0 ectodomains in solution. We then generated a 24-Å-resolution 3D reconstruction. The NTD-G2 Fab crystal structure was superimposed onto the prefusion MERS-CoV S structure (PDB: <ext-link ext-link-type="uri" xlink:href="pdb:5W9J" id="intref0010">5W9J</ext-link>) to generate a model, which fit well into the reconstruction (<xref rid="mmc1" ref-type="supplementary-material">Figure S3</xref>B), indicating that G2 Fab binding does not induce substantial conformational changes in the MERS-CoV S trimer. Note that density for the RBD is missing due to the intrinsic dynamics of the RBD (<xref rid="bib19" ref-type="bibr">Gui et al., 2017</xref>, <xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>, <xref rid="bib74" ref-type="bibr">Yuan et al., 2017</xref>).</p><p id="p0110">To further investigate the interaction of G2 with the MERS-CoV S trimer, we determined a 4.2-Å cryo-EM structure of a MERS-CoV S0-G2 Fab complex (<xref rid="fig3" ref-type="fig">Figures 3</xref>A, <xref rid="mmc1" ref-type="supplementary-material">S3</xref>C, and S3D; <xref rid="mmc1" ref-type="supplementary-material">Table S3</xref>). The MERS-CoV S1 NTDs reside on the periphery of the S1 trimer, flared out approximately 45° to the 3-fold axis. The G2 interface is situated at the apex of the S protein, and binding of G2 Fab elongates the S1-NTD axis to form three legs of an inverted “tripod” structure, with a 15° tilt toward the 3-fold axis (<xref rid="fig3" ref-type="fig">Figure 3</xref>B). The distance between two bound Fabs is 110 Å at the binding interface and 145 Å at the distal end of the Fab (<xref rid="fig3" ref-type="fig">Figure 3</xref>A). There are no substantial conformational changes in the trimer resulting from G2 binding.<fig id="fig3"><label>Figure 3</label><caption><p>G2 Binding to the Prefusion MERS-CoV Spike</p><p>(A) Cryo-EM structure of uncleaved MERS-CoV S0 ectodomain in complex with G2 Fab as viewed along the viral membrane. A single protomer of the trimeric S protein is shown in ribbon representation, with S1-NTD and G2 colored the same as in <xref rid="fig2" ref-type="fig">Figure 2</xref> and the rest of S colored light blue. See also <xref rid="mmc1" ref-type="supplementary-material">Figures S3</xref>C and S3D.</p><p>(B) Structural model of G2 Fab bound to one MERS-CoV S protomer. The angles of the S1-NTD and G2 Fab to the 3-fold axis are depicted.</p></caption><graphic xlink:href="gr3_lrg"></graphic></fig></p></sec><sec id="sec2.5"><title>G2 Prevents S Protein Binding to the Cell Surface</title><p id="p0115">The binding of G2 Fab to the apex of the spike, where interactions with host-cell factors such as DPP4 occur, suggested that G2 IgG may interfere with the attachment step during cell entry. To test this hypothesis, we performed cell-surface binding assays. As shown in <xref rid="fig4" ref-type="fig">Figure 4</xref>, GFP-labeled MERS-CoV S-2P strongly bound to DPP4-transfected HEK293 cells. This binding was substantially reduced by pre-incubation of MERS-CoV S-2P with a 5-fold molar excess of G2 IgG or an RBD-specific antibody, D12. Interestingly, G2 reduced MERS-CoV S-2P binding more than D12 did, even though they have similar neutralizing potencies (<xref rid="bib68" ref-type="bibr">Wang et al., 2015</xref>). MERS-CoV S-2P variants with substitutions S28F or G198D bound to cells at levels similar to those observed for WT MERS-CoV S-2P. However, in contrast to WT MERS-CoV S-2P, these two variants were insensitive to G2 IgG and maintained high-level cell-surface binding even in the presence of excess G2 IgG. Collectively, these data demonstrate that G2 IgG prevents attachment of MERS-CoV S to cells expressing DPP4.<fig id="fig4"><label>Figure 4</label><caption><p>G2 IgG Prevents the Binding of MERS-CoV S Protein to DDP4-Expressing Cells</p><p>Normalized binding efficiency of GFP-tagged MERS-CoV S-2P proteins to DPP4-expressing FreeStyle 293F cells in the presence or absence of IgGs was calculated from median fluorescence intensity (MFI) values. FreeStyle 293-F cells were transfected with a plasmid encoding full-length DPP4 60 h before the experiment. Non-transfected cells (NTs) incubated with MERS-CoV S-2P, as well as transfected cells incubated with PBS, were used as negative controls. AM14 is an irrelevant RSV F-specific neutralizing antibody used as another negative control. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates).</p></caption><graphic xlink:href="gr4_lrg"></graphic></fig></p></sec><sec id="sec2.6"><title>G2 Fab Neutralizes but Not through a Direct Steric Clash with DPP4</title><p id="p0120">The binding of MERS-CoV S to DPP4 requires the RBDs to rotate upward to expose the receptor binding surface (<xref rid="fig5" ref-type="fig">Figure 5</xref>A). Presumably, antibodies can block the receptor-binding process by inhibiting RBD movement or DPP4 attachment. To gain insight into the mechanism of G2 attachment inhibition, we superimposed G2 and DPP4 onto the MERS-CoV S structure using the S1-NTD-G2 Fab structure presented here and the previously determined DPP4-RBD structure (<xref rid="bib35" ref-type="bibr">Lu et al., 2013</xref>, <xref rid="bib70" ref-type="bibr">Wang et al., 2013</xref>). Surprisingly, the structural model indicates that G2 Fab would not prevent movement of the RBD nor sterically clash with DPP4 (<xref rid="fig5" ref-type="fig">Figure 5</xref>B). However, a steric clash would be predicted to occur between MERS-CoV S and the host-cell membrane if G2 IgG were bound (<xref rid="fig5" ref-type="fig">Figure 5</xref>C), and the bivalent G2 IgG could also possibly cross-link two adjacent MERS-CoV S trimers, resulting in restricted access for DPP4.<fig id="fig5"><label>Figure 5</label><caption><p>Comparison of DPP4 Binding-Inhibition and Neutralization Activity of G2 IgG versus Fab</p><p>(A–C) Structural models of MERS-CoV S trimers with a single RBD in the up conformation (A) unbound, (B) bound to G2 Fab and DPP4, and (C) bound to G2 IgG. Models were generated in PyMOL based on superimposed structures of MERS-CoV S (PDB: <ext-link ext-link-type="uri" xlink:href="pdb:5W9H" id="intref0180">5W9H</ext-link>), RBD-DPP4 complex (PDB: <ext-link ext-link-type="uri" xlink:href="pdb:4L72" id="intref0185">4L72</ext-link>), mouse IgG1 (PDB: <ext-link ext-link-type="uri" xlink:href="pdb:1IGY" id="intref0190">1IGY</ext-link>), and the S1-NTD–G2 structure described in this paper. MERS-CoV S protomers are colored green, pink, and orange, with the green protomer in the “RBD up” conformation. The DPP4 dimer is colored red, whereas the G2 heavy and light chains are colored blue and white, respectively.</p><p>(D) Inhibition of soluble MERS-CoV S-2P binding to DDP4-expressing cells as a function of IgG or Fab concentration. The mean of duplicate measurements is plotted. Error bars represent SEM.</p><p>(E) Neutralization of MERS-CoV pseudoviruses as a function of IgG or Fab concentration. The mean of duplicate measurements is plotted. Error bars represent SEM.</p><p>(F) Neutralization of authentic MERS-CoV as a function of IgG or Fab concentration. The mean of duplicate measurements is plotted. The IgG data are the same as those plotted in <xref rid="fig1" ref-type="fig">Figure 1</xref>F.</p></caption><graphic xlink:href="gr5_lrg"></graphic></fig></p><p id="p0125">To determine whether the G2 Fab, rather than the larger IgG, was sufficient for activity, the binding inhibition and neutralizing ability of G2 Fab were evaluated. Consistent with our previous data (<xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>, <xref rid="bib53" ref-type="bibr">Rosen et al., 2019</xref>), G2 IgG displayed strong inhibition of soluble MERS-CoV S binding to DPP4-expressing BHK21 cells with an IC<sub>50</sub> of 15 nM (<xref rid="fig5" ref-type="fig">Figure 5</xref>D), as well as strong neutralization of MERS-CoV S-containing pseudoviruses with an IC<sub>50</sub> of 0.09 nM (<xref rid="fig5" ref-type="fig">Figure 5</xref>E). G2 Fab also inhibited cell binding (IC<sub>50</sub> = 32.3 nM) (<xref rid="fig5" ref-type="fig">Figure 5</xref>D) and neutralized MERS-CoV S-containing pseudoviruses (IC<sub>50</sub> = 1.33 nM), although not as well as G2 IgG (<xref rid="fig5" ref-type="fig">Figure 5</xref>E). G2 Fab also neutralized authentic MERS-CoV in a dose-dependent manner, but as we observed with the pseudoviruses, the neutralizing activity of the Fab was not as strong as the IgG (<xref rid="fig5" ref-type="fig">Figure 5</xref>F). Based on these data, we conclude that G2 Fab is sufficient for neutralization and blocking attachment of MERS-CoV S to DPP4-expressing cells, but likely not by clashing with DPP4 nor restricting exposure of the RBDs.</p></sec><sec id="sec2.7"><title>G2 Neutralizes by Preventing the Binding of Trimeric S Protein with DPP4</title><p id="p0130">Although it is possible that the S1-NTD contributes to the cell-surface attachment process by directly binding one or more attachment factors, the RBD plays the predominant role in this event by strongly binding to the functional receptor DPP4. We hypothesized that G2 blocks cell-surface attachment by indirectly inhibiting the RBD-DPP4 interaction. To test this, we investigated cell-surface binding of soluble DPP4 to membrane-anchored MERS-CoV S1 (S1-TM), full-length S-WT (S-WT-FL), or full-length S-2P (S-2P-FL) in the presence or absence of excess G2 Fab. We observed that DPP4 bound well to cells transfected with each of the three S protein constructs (<xref rid="fig6" ref-type="fig">Figures 6</xref>A–6C), as expected. As a positive antibody control, RBD-specific JC57-14 Fab strongly inhibited binding of DPP4 to each of the transfected cells by directly competing with DPP4 for RBD binding (<xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>). As a negative antibody control, the S2-specific G4 Fab (<xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>, <xref rid="bib68" ref-type="bibr">Wang et al., 2015</xref>) exhibited no binding inhibition. When G2 Fab was tested, a different binding-inhibition pattern was observed. G2 Fab prevented binding of DPP4 to cells expressing trimeric full-length S-WT or S-2P, but allowed binding of DPP4 to cells expressing a membrane-tethered form of S1 that is predicted to be monomeric (<xref rid="fig6" ref-type="fig">Figures 6</xref>A–6C). These data indicate that DPP4 binding inhibition by G2 Fab depends on the intact trimeric conformation of the S protein.<fig id="fig6"><label>Figure 6</label><caption><p>G2-Mediated Inhibition of DPP4 Binding to MERS-CoV S Depends on the Oligomeric State of the Spike</p><p>(A–C) Normalized binding efficiency of DPP4, in the presence or absence of Fab, to cells transfected with plasmids encoding (A) membrane-anchored S1 (S1-TM), (B) full-length S-WT (S-WT-FL), and (C) full-length S-2P (S-2P-FL). Cells incubated with PBS were used as negative controls. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates).</p><p>(D) Surface plasmon resonance competition assay. Response curves for MERS-CoV S-2P, alone or in the presence of a 5-fold molar excess of indicated Fabs or IgGs, passed over an immobilized DPP4 ectodomain. The curves for S-2P or S-2P supplemented with Fab or IgG are shown with a solid line, whereas control curves for samples without S-2P are shown with a dotted line.</p></caption><graphic xlink:href="gr6_lrg"></graphic></fig></p><p id="p0135">To exclude the possibility that unknown cell-surface factors may play roles in G2’s inhibition of DPP4 binding to MERS-CoV S trimers, we performed an SPR competition assay using purified proteins (<xref rid="fig6" ref-type="fig">Figure 6</xref>D). We captured DPP4 onto the SPR chip and then flowed over MERS-CoV S-2P protein or MERS-CoV S-2P protein supplemented with a 5-fold molar excess of different Fabs. MERS-CoV S-2P in the absence of Fabs bound well to the captured DPP4 (black curve). MERS-CoV S-2P supplemented with the RBD-specific JC57-14 Fab displayed no binding (blue curve), confirming the strong competition between JC57-14 Fab and DPP4 for binding to the RBD. MERS-CoV S-2P supplemented with the S2-specific G4 Fab displayed a higher response curve than MERS-CoV S-2P alone due to the added mass of the bound G4 Fab and its lack of binding inhibition. MERS-CoV S-2P supplemented with G2 Fab displayed substantially lower binding to the captured DPP4 than either the MERS-CoV S-2P supplemented with G4 Fab or MERS-CoV S2-P alone (purple curve), confirming that G2 interferes with DPP4 binding to trimeric MERS-CoV S. G2 IgG reduced the binding of MERS-CoV S-2P even further (red curve), despite the increased mass of the IgG compared to the Fab. Unlike JC57-14 Fab, neither G2 Fab nor G2 IgG blocked DPP4 binding completely, suggesting that G2 uses an indirect mechanism of inhibition.</p></sec></sec><sec id="sec3"><title>Discussion</title><p id="p0140">Around 20 different monoclonal antibodies have been isolated that can neutralize MERS-CoV, with the majority targeting the RBD. With the exception of the S2-specific antibody G4, the other non-RBD-directed MERS-CoV neutralizing antibodies that have been isolated are S1-NTD specific, including G2 and 5F9 from mice, JC57-13 and FIB-H1 from macaques, and CDC2-A2 from a human (<xref rid="bib7" ref-type="bibr">Chen et al., 2017</xref>, <xref rid="bib68" ref-type="bibr">Wang et al., 2015</xref>, <xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>). S1-NTD-specific neutralizing antibodies have also been isolated against SARS-CoV (<xref rid="bib11" ref-type="bibr">Coughlin et al., 2009</xref>, <xref rid="bib18" ref-type="bibr">Greenough et al., 2005</xref>, <xref rid="bib52" ref-type="bibr">Rockx et al., 2008</xref>), including 4D4, 68, S132, and S228.11. Thus, for SARS-CoV and MERS-CoV, the S1-NTD is a common site of antibody vulnerability. Despite a lack of structural characterization, studies with these antibodies provide some insight into their neutralizing mechanism. Antibody 4D4 was shown to bind the SARS S1-NTD fragment between residues 12 and 261 (<xref rid="bib11" ref-type="bibr">Coughlin et al., 2009</xref>). 4D4 efficiently prevented viral entry when added after the binding of pseudovirus onto target cells, suggesting 4D4 is more likely to neutralize through disrupting a post-binding event instead of interfering with the cell attachment process. We also note that another MERS-CoV S1-NTD-specific neutralizing antibody, A2, does not compete with G2 (<xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>), indicating that G2 and A2 recognize different epitopes on the S1-NTD. Thus, it is likely that S1-NTD-derived antibodies can neutralize through different mechanisms.</p><p id="p0145">The molecular mechanisms regulating receptor binding and fusion activation of coronavirus spike proteins are currently the subject of active investigation in the field. Recent studies have demonstrated that both receptor binding and proteolytic cleavage are required to shed S1 and allow refolding of S2 to the postfusion state (<xref rid="bib58" ref-type="bibr">Song et al., 2018</xref>, <xref rid="bib67" ref-type="bibr">Walls et al., 2019</xref>). Our data suggest that the S1-NTD may play a role in regulating this finely tuned process, although the molecular details remain unknown. Upon binding the NTDs, the large bivalent G2 IgG would be able to generate a barrier at the virus-host interface to prevent attachment (<xref rid="fig5" ref-type="fig">Figure 5</xref>C). DPP4 has a short 11-residue stalk, which restricts its movement and access to the RBDs of a G2-bound S protein. This can reasonably explain the neutralization ability exerted by G2 IgG. However, we demonstrated that G2 Fab can also prevent the binding of MERS-CoV S trimer with DPP4 despite no predicted steric clash between the G2 Fab and cell-surface DPP4 (<xref rid="fig5" ref-type="fig">Figure 5</xref>B). We also demonstrated that G2 Fab has the ability to prevent the attachment of soluble DPP4 to cell-surface S trimer (<xref rid="fig6" ref-type="fig">Figure 6</xref>). We feel it is unlikely that the reduced DPP4 binding can be attributed to steric interference with G2. It may be that there is some cross talk between the S1-NTDs and RBDs within the trimer, such that G2 binding to the NTDs reduces exposure of the RBDs in the receptor-accessible “up” conformation. A previous study identified a SARS-CoV S1-NTD-derived peptide (residues 217-234), designated “peptide 9626,” that inhibits SARS-CoV S-mediated entry in a dose-dependent manner (<xref rid="bib20" ref-type="bibr">Guo et al., 2009</xref>). When this peptide is mapped onto the cryo-EM structure of the SARS-CoV S trimer, it localizes to the S1-NTD-RBD interface, suggesting that the inhibition is mediated by interfering with the interaction between an S1-NTD and an RBD from an adjacent protomer.</p><p id="p0150">Our data from the <italic>in vitro</italic> competition assay demonstrated that G2 Fab indirectly prevents the RBD-DPP4 interaction in the absence of any other attachment factors or co-receptors. However, several MERS-CoV attachment factors have been identified, including CEACAM5 (<xref rid="bib6" ref-type="bibr">Chan et al., 2016</xref>), sialic acid (<xref rid="bib33" ref-type="bibr">Li et al., 2017</xref>), and GRP78 (<xref rid="bib8" ref-type="bibr">Chu et al., 2018</xref>). Several coronaviruses have been demonstrated to use the S1-NTD to bind glycans and the S1-CTD to bind the functional protein receptor (<xref rid="bib29" ref-type="bibr">Li, 2015</xref>). In the case of TGEV, a porcine coronavirus, S1-NTD-mediated sialic acid binding is highly related to enteropathogenicity (<xref rid="bib25" ref-type="bibr">Krempl et al., 1997</xref>). BCoV (<xref rid="bib46" ref-type="bibr">Peng et al., 2012</xref>, <xref rid="bib54" ref-type="bibr">Schultze et al., 1991</xref>) and OC43 (<xref rid="bib27" ref-type="bibr">Künkel and Herrler, 1993</xref>) also use sialic acid as a receptor, with the sugar-binding pocket located on the S1-NTD top region. MHV, another β-coronavirus, uses the S1-NTD top region to bind its protein receptor CEACAM1 (<xref rid="bib45" ref-type="bibr">Peng et al., 2011</xref>). Interestingly, although BCoV and MHV spikes bind to different host receptors, their S1-NTD binding surfaces are similarly presented (<xref rid="fig7" ref-type="fig">Figure 7</xref>), with the loop stretching out from β5 to β6 contributing most of the binding surface. G2-like antibodies may therefore directly block these receptor interactions for other coronaviruses.<fig id="fig7"><label>Figure 7</label><caption><p>Comparison of the S1-NTD Top Region from MERS-CoV, BCoV, and MHV</p><p>S1-NTD is colored as in <xref rid="fig2" ref-type="fig">Figure 2</xref> and depicted in ribbon representation in the top and middle rows. Residues interacting with G2 Fab, sialic acid, and CEACAM1 are shown as blue spheres on the MERS-CoV, BCoV, and MHV structures, respectively. Topology models of the S1-NTDs are shown in the bottom row, with β sheets depicted as arrows and α helices depicted as red cylinders. The binding surfaces described above are denoted with a blue oval. Figures were made based on the structure of MERS-CoV S1-NTD–G2 Fab described in this paper, BCoV S1-NTD (<xref rid="bib46" ref-type="bibr">Peng et al., 2012</xref>) (PDB: <ext-link ext-link-type="uri" xlink:href="pdb:4H14" id="intref0195">4H14</ext-link>), and MHV S (<xref rid="bib45" ref-type="bibr">Peng et al., 2011</xref>) (PDB: <ext-link ext-link-type="uri" xlink:href="pdb:3JCL" id="intref0200">3JCL</ext-link>).</p></caption><graphic xlink:href="gr7_lrg"></graphic></fig></p><p id="p0155">As mentioned above, numerous S1-NTD-specific neutralizing antibodies have been isolated from mice, non-human primates (NHPs), and MERS-CoV patients (<xref rid="bib7" ref-type="bibr">Chen et al., 2017</xref>, <xref rid="bib68" ref-type="bibr">Wang et al., 2015</xref>, <xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>), indicating that the S1-NTD is immunogenic. A recent study tested this directly by using a MERS-CoV S1-NTD fragment as an immunogen, and the results revealed that the S1-NTD can induce robust protective antibody responses in a mouse model (<xref rid="bib21" ref-type="bibr">Jiaming et al., 2017</xref>). Our study defines a site of vulnerability on the S1-NTD, which may be important for structure-based vaccine design. Future efforts may improve the immunogenicity of the S1-NTD by exposing the vulnerable sites while masking or eliminating non-neutralizing epitopes, such as those normally buried in the prefusion spike. This strategy has already been successfully used for RBD-based vaccine design (<xref rid="bib13" ref-type="bibr">Du et al., 2016</xref>), wherein the introduction of glycosylation sites in the non-neutralizing epitopes led to an engineered RBD immunogen that was significantly more efficacious in a mouse model of MERS-CoV challenge.</p></sec><sec id="sec4"><title>STAR★Methods</title><sec id="sec4.1"><title>Key Resources Table</title><p id="p0160"><table-wrap position="float" id="undtbl1"><table frame="hsides" rules="groups"><thead><tr><th>REAGENT or RESOURCE</th><th>SOURCE</th><th>IDENTIFIER</th></tr></thead><tbody><tr><td colspan="3"><bold>Antibodies</bold></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td>G2</td><td><xref rid="bib68" ref-type="bibr">Wang et al., 2015</xref></td><td>N/A</td></tr><tr><td>G4</td><td><xref rid="bib68" ref-type="bibr">Wang et al., 2015</xref></td><td>N/A</td></tr><tr><td>D12</td><td><xref rid="bib68" ref-type="bibr">Wang et al., 2015</xref></td><td>N/A</td></tr><tr><td>JC57-14</td><td><xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref></td><td>N/A</td></tr><tr><td>Goat anti-human IgG, Alexa Fluor ® 647-conjugated</td><td>ThermoFisher Scientific</td><td>Cat#A-21445</td></tr><tr><td>Goat anti-rabbit IgG, Alexa Fluor® 488-conjugated</td><td>Abcam</td><td>Cat#ab150089</td></tr><tr><td>MERS-CoV S Antibody, Rabbit pAb</td><td>Sino Biological</td><td>Cat#40069</td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td colspan="3"><bold>Bacterial and Virus Strains</bold></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td>MERS-CoV EMC/2012</td><td><xref rid="bib55" ref-type="bibr">Scobey et al., 2013</xref></td><td>N/A</td></tr><tr><td>MERS-CoV EMC/2012</td><td><xref rid="bib2" ref-type="bibr">Almazán et al., 2013</xref></td><td>N/A</td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td colspan="3"><bold>Chemicals, Peptides, and Recombinant Proteins</bold></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td>Kifunensine</td><td>GlycoSyn</td><td>Cat#FC-034</td></tr><tr><td>DAPI</td><td>ThermoFisher Scientific</td><td>Cat#D1306</td></tr><tr><td>25 kDa linear polyethylenimine</td><td>Polysciences</td><td>Cat#3966-2</td></tr><tr><td>Fugene 6 transfection reagent</td><td>Promega</td><td>Cat#E2691</td></tr><tr><td>Lipofectamine 3000 reagent</td><td>ThermoFisher Scientific</td><td>Cat#L3000001</td></tr><tr><td>TRIzol reagent</td><td>ThermoFisher Scientific</td><td>Cat#15596026</td></tr><tr><td>MERS-CoV S-2P</td><td><xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref></td><td>N/A</td></tr><tr><td>MERS-CoV S0</td><td>This paper</td><td>N/A</td></tr><tr><td>MERS-CoV S-2P-GFP</td><td>This paper</td><td>N/A</td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td colspan="3"><bold>Critical Commercial Assays</bold></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td>Luciferase assay kit</td><td>Promega</td><td>Cat#E1501</td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td colspan="3"><bold>Deposited Data</bold></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td>G2 Fab structure</td><td>This paper</td><td>PDB: <ext-link ext-link-type="uri" xlink:href="pdb:6PXG" id="intref0015">6PXG</ext-link></td></tr><tr><td>MERS-CoV S1-NTD–G2 Fab structure</td><td>This paper</td><td>PDB: <ext-link ext-link-type="uri" xlink:href="pdb:6PXH" id="intref0020">6PXH</ext-link></td></tr><tr><td>Cryo-EM map of MERS-S0–G2 Fab complex</td><td>This paper</td><td>EMDB: EMD-20527</td></tr><tr><td>Coordinates for MERS-S0–G2 Fab complex</td><td>This paper</td><td>PDB: <ext-link ext-link-type="uri" xlink:href="pdb:6PZ8" id="intref0025">6PZ8</ext-link></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td colspan="3"><bold>Experimental Models: Cell Lines</bold></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td>FreeStyle 293-F Cells</td><td>ThermoFisher Scientific</td><td>Cat#R79007</td></tr><tr><td>Expi293F Cells</td><td>ThermoFisher Scientific</td><td>Cat#A14527</td></tr><tr><td>Vero 81 cells</td><td>ATCC</td><td>CCL-81</td></tr><tr><td>293T cells</td><td>ATCC</td><td>CRL-11268</td></tr><tr><td>BHK-21 cells</td><td>ATCC</td><td>CCL-10 TM</td></tr><tr><td>Huh7.5 cells</td><td>Provided by Dr. Deborah R. Taylor of the US FDA</td><td>N/A</td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td colspan="3"><bold>Recombinant DNA</bold></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td>pCMVDR8.2</td><td><xref rid="bib59" ref-type="bibr">Stewart et al., 2003</xref></td><td>Addgene Cat#8455</td></tr><tr><td>pHR′ CMV-Luc</td><td>Barney Graham Laboratory</td><td>N/A</td></tr><tr><td>CMV/R-MERS-CoV S</td><td>Barney Graham Laboratory</td><td>N/A</td></tr><tr><td>pαH expression plasmid</td><td>Jason McLellan Laboratory</td><td>N/A</td></tr><tr><td>pVRC8400 expression plasmid</td><td>Barney Graham Laboratory</td><td>N/A</td></tr><tr><td>pαH-S1-TM</td><td>This paper</td><td>N/A</td></tr><tr><td>pαH-S-WT-FL</td><td>This paper</td><td>N/A</td></tr><tr><td>pαH-S-2P-FL</td><td>This paper</td><td>N/A</td></tr><tr><td>pαH-DPP4</td><td>This paper</td><td>N/A</td></tr><tr><td>paH-DPP4-ECD-HSS</td><td>This paper</td><td>N/A</td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td colspan="3"><bold>Software and Algorithms</bold></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td>Prism (V7)</td><td>GraphPad</td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_002798" id="intref0030">SCR_002798</ext-link></td></tr><tr><td>FlowJo (V7.6.1)</td><td>FlowJo</td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_008520" id="intref0035">SCR_008520</ext-link></td></tr><tr><td>Scrubber2</td><td>BioLogic</td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_015745" id="intref0040">SCR_015745</ext-link></td></tr><tr><td>cryoSPARC v2</td><td><xref rid="bib50" ref-type="bibr">Punjani et al., 2017</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_016501" id="intref0045">SCR_016501</ext-link></td></tr><tr><td>Pymol</td><td>Schrödinger</td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_000305" id="intref0050">SCR_000305</ext-link></td></tr><tr><td>UCSF Chimera</td><td><xref rid="bib47" ref-type="bibr">Pettersen et al., 2004</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_004097" id="intref0055">SCR_004097</ext-link></td></tr><tr><td>CCP4i interface</td><td><xref rid="bib49" ref-type="bibr">Potterton et al., 2003</xref></td><td><ext-link ext-link-type="uri" xlink:href="http://www.ccp4.ac.uk" id="intref0060">http://www.ccp4.ac.uk</ext-link></td></tr><tr><td>PHASER</td><td><xref rid="bib36" ref-type="bibr">McCoy et al., 2007</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_014219" id="intref0065">SCR_014219</ext-link></td></tr><tr><td>COOT</td><td><xref rid="bib14" ref-type="bibr">Emsley and Cowtan, 2004</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_014222" id="intref0070">SCR_014222</ext-link></td></tr><tr><td>Phenix</td><td><xref rid="bib1" ref-type="bibr">Adams et al., 2002</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_014224" id="intref0075">SCR_014224</ext-link></td></tr><tr><td>Leginon software suite</td><td><xref rid="bib48" ref-type="bibr">Potter et al., 1999</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_016731" id="intref0080">SCR_016731</ext-link></td></tr><tr><td>Appion</td><td><xref rid="bib28" ref-type="bibr">Lander et al., 2009</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_016734" id="intref0085">SCR_016734</ext-link></td></tr><tr><td>EMAN2</td><td><xref rid="bib61" ref-type="bibr">Tang et al., 2007</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_016867" id="intref0090">SCR_016867</ext-link></td></tr><tr><td>Rosetta (v2019.14.60699)</td><td><xref rid="bib9" ref-type="bibr">Conway et al., 2014</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_015701" id="intref0095">SCR_015701</ext-link></td></tr><tr><td>PISA</td><td><xref rid="bib26" ref-type="bibr">Krissinel and Henrick, 2007</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_015749" id="intref0100">SCR_015749</ext-link></td></tr><tr><td>MotionCor2</td><td><xref rid="bib78" ref-type="bibr">Zheng et al., 2017</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_016499" id="intref0105">SCR_016499</ext-link></td></tr><tr><td>GCTF</td><td><xref rid="bib77" ref-type="bibr">Zhang, 2016</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_016500" id="intref0110">SCR_016500</ext-link></td></tr><tr><td>DoG Picker</td><td><xref rid="bib64" ref-type="bibr">Voss et al., 2009</xref></td><td>RRID: <ext-link ext-link-type="uri" xlink:href="rridsoftware:SCR_016655" id="intref0115">SCR_016655</ext-link></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td colspan="3"><bold>Other</bold></td></tr><tr><td colspan="3"><hr></hr></td></tr><tr><td>Strep-Tactin Superflow resin</td><td>IBA Lifesciences</td><td>Cat# 2-1206-010</td></tr><tr><td>Biacore Sensor Chip NTA</td><td>GE Healthcare</td><td>Cat#BR100407</td></tr><tr><td>Biacore Sensor Chip NTA CM5</td><td>GE Healthcare</td><td>Cat# BR100399</td></tr><tr><td>Protein A agarose</td><td>ThermoFisher Scientific</td><td>Cat#20334</td></tr><tr><td>HiLoad 16/600 Superdex 75 pg column</td><td>GE Healthcare</td><td>Cat#28989333</td></tr><tr><td>HiLoad 16/600 Superdex 200 pg column</td><td>GE Healthcare</td><td>Cat#28989335</td></tr></tbody></table></table-wrap></p></sec><sec id="sec4.2"><title>Lead Contact and Materials Availability</title><p id="p0165">Further information and requests for resources and reagents should be directed to and will be fulfilled by the Lead Contact, Jason S. McLellan (<ext-link ext-link-type="uri" xlink:href="mailto:jmclellan@austin.utexas.edu" id="intref0120">jmclellan@austin.utexas.edu</ext-link>). Reagents generated in this study are available via material transfer agreement (MTA).</p></sec><sec id="sec4.3"><title>Experimental Model and Subject Details</title><sec id="sec4.3.1"><title>Cell Lines</title><p id="p0170">FreeStyle 293F cells and Expi293 cells were purchased from Thermofisher Scientific and used to express proteins, Fabs and IgGs. They are maintained following the company’s protocol.</p><p id="p0175">Vero 81 cells were obtained from ATCC, cultured in Dulbecco’s Modified Eagle Medium (supplemented with 10% fetal bovine serum, 100 U/ml penicillin, 100 μg/ml streptomycin, and 0.25 μg/ml amphotericin B) in a humidified 37 °C incubator containing 5% CO<sub>2</sub>, and used for <italic>in vitro</italic> selection of G2-escape variants and plaque-reduction neutralization assay.</p><p id="p0180">HEK293T cells were obtained from ATCC, cultured in DMEM with 10% heat-inactivated fetal bovine serum (HI-FBS) and 1 × penicillin/streptomycin in a 37 °C incubator containing 5% CO<sub>2</sub>, and used to produce MERS-CoV pseudoviruses.</p><p id="p0185">BHK-21 cells were obtained from ATCC, cultured in DMEM medium with 10% HI-FBS and 1 × penicillin/streptomycin in a 37 °C incubator containing 5% CO<sub>2</sub>.</p><p id="p0190">Huh7.5 cells were provided by Dr Deborah R. Taylor of the US FDA and cultured in DMEM with 10% HI-FBS and 1 × penicillin/streptomycin in a 37 °C incubator containing 5% CO<sub>2</sub>.</p><p id="p0195">BHK-21 and Huh7.5 cells were used for neutralization assays in different experiments.</p></sec><sec id="sec4.3.2"><title>Viral Strains</title><p id="p0200">Recombinant MERS-CoV strain EMC/2012 (<xref rid="bib55" ref-type="bibr">Scobey et al., 2013</xref>) was used for selection of G2-escape variants. For plaque-reduction neutralization testing, recombinant wild-type and spike mutant MERS-CoV were recovered from a bacterial artificial chromosome (BAC) containing the full-length EMC/2012 isolate genome (<xref rid="bib2" ref-type="bibr">Almazán et al., 2013</xref>), a gift from Dr. Stanley Perlman (University of Iowa). The G198D spike mutation, which arose during passage selection of MERS-CoV for resistance to G2-mediated neutralization, was introduced into the MERS-CoV BAC using published protocols (<xref rid="bib16" ref-type="bibr">Fehr et al., 2015</xref>). The complete S gene sequence of recombinant virus was determined to confirm accuracy of mutagenesis.</p></sec></sec><sec id="sec4.4"><title>Method Details</title><sec id="sec4.4.1"><title>Selection and Analysis of G2-Escape Variants</title><p id="p0205">A P0 stock of recombinant MERS-CoV EMC/2012 recovered from an infectious clone (<xref rid="bib55" ref-type="bibr">Scobey et al., 2013</xref>) was serially passaged in Vero 81 cells supplemented with increasing concentrations of G2. Three parallel passage series were performed. Passages were initiated at a multiplicity of infection (MOI) ∼0.001 PFU per cell and G2 concentration = 0.43 μg/ml, corresponding to 0.75x G2 IC80 versus MERS-CoV in a plaque-reduction neutralization assay. Culture supernatants were passed onto fresh cells when 50%–60% of the monolayer displayed viral CPE. Viral inoculum volume and G2 concentrations were empirically co-adjusted between passage steps to produce the target CPE level at approximately 48 hours-post-infection (hpi). The G2 concentration at terminal passage, P10, was 0.65 μg/ml, corresponding to 11.2x IC80. 15 clonal escape mutant viruses (five from each passage) were isolated from P10 cultures via plaque purification on Vero 81 cells in the presence of 0.65 μg/ml G2. Viral plaques were expanded in Vero 81 cells, and total RNA was harvested in TRIzol reagent (Invitrogen), followed by RT-PCR to generate overlapping cDNA amplicons spanning the entire S gene open reading frame. PCR products were subjected to dideoxy sequencing, and reads were aligned to the native EMC/2012 S gene sequence (GenBank accession number <ext-link ext-link-type="uri" xlink:href="ncbi-n:JX869059.2" id="intref0125">JX869059.2</ext-link>) to identify differences. In a separate virus passage experiment, three parallel lineages of antibody-free P10 EMC/2012 cultures were examined for S mutations to identify cell culture-adaptive changes resulting from serial MERS-CoV passage. These substitutions were excluded from analysis of changes in S identified in G2 escape mutant isolates.</p></sec><sec id="sec4.4.2"><title>Plaque-Reduction Neutralization Assay</title><p id="p0210">Wild-type and spike mutant MERS-CoV were recovered from a bacterial artificial chromosome (BAC) containing the full-length EMC/2012 isolate genome (<xref rid="bib2" ref-type="bibr">Almazán et al., 2013</xref>), a gift from Dr. Stanley Perlman (University of Iowa). The G198D spike mutation, which arose during passage selection of MERS-CoV for resistance to G2-mediated neutralization, was introduced into the MERS-CoV BAC using published protocols (<xref rid="bib16" ref-type="bibr">Fehr et al., 2015</xref>). The complete S gene sequence of recombinant virus was determined to confirm accuracy of mutagenesis.</p><p id="p0215">Serial 2- or 10-fold dilutions of G2 IgG or Fab were combined with approximately 70–130 PFU of WT or G198D mutant MERS-CoV in a total volume of 200 μL gelatin saline (0.3% [wt/vol] gelatin in phosphate-buffered saline supplemented with CaCl<sub>2</sub> and MgCl<sub>2</sub>). Virus-antibody mixtures were incubated for 20 min at 37°C, followed by adsorption of 100 μL aliquots to each of two confluent wells of Vero 81 cells in 6-well (10-cm<sup>2</sup>) plates for 30 min at 37°C. Monolayers were overlaid with Dulbecco’s modified Eagle’s medium (DMEM) containing 1% agar, and plaques were enumerated at 96 h post-infection. Percent neutralization was calculated as average plaques produced by IgG- or Fab-treated virus divided by average plaques produced by virus in antibody-free gelatin saline.</p></sec><sec id="sec4.4.3"><title>Pseudovirus Infectivity Experiments</title><p id="p0220">The S28F and G198D substitutions were introduced via PCR into a plasmid encoding full-length spike (MERS-CoV England1 AFY 13307). S-containing lentiviral pseudovirions (Eng1, Eng1-S28F and Eng1-G198D) were produced by co-transfection of three plasmids (packaging plasmid pCMVDR8.2, transducing plasmid pHR′ CMV-Luc, and CMV/R-MERS-CoV S plasmid) into 293T cells using Fugene 6 transfection reagent (Promega, Madison, WI) (<xref rid="bib41" ref-type="bibr">Naldini et al., 1996</xref>). For the neutralization assay, Huh7.5 cells, provided by Deborah R. Taylor of the U.S. FDA, were plated into 96-well white/black Isoplates (PerkinElmer, Waltham, MA) at 10,000 cells/well the day before infection. Serial dilutions of monoclonal antibody were mixed with titrated pseudovirus, incubated for 30 min at 37°C, and added to Huh7.5 cells in triplicate. Following 2 h of incubation, wells were replenished with 100 μL of fresh medium. Cells were lysed 72 h later, and luciferase activity was measured. Percent neutralization was calculated from luminometry data.</p></sec><sec id="sec4.4.4"><title>Production of G2 Fab</title><p id="p0225">The Fab region of the G2 heavy chain was fused with the HRV3C cleavage site followed by human IgG1 Fc fragment and subcloned into the eukaryotic expression vector pVRC8400. This plasmid (plasmid 1) was cotransfected with a vector encoding the G2 light chain (plasmid 2) into Expi293 cells (Invitrogen), and the secreted antibody was purified using Protein A agarose (Fisher). HRV3C protease (1% wt/wt) was added to the protein and the reaction was incubated for 2 h at room temperature. The digested antibody was passed back over Protein A agarose to remove the Fc fragment, and the unbound Fab was additionally purified using a Superdex 75 column (GE Healthcare).</p></sec><sec id="sec4.4.5"><title>Production of MERS-CoV S1-NTD and Mutants</title><p id="p0230">A gene encoding MERS-CoV S1-NTD (residues 1–351) with a C-terminal HRV3C cleavage site, 8xHis-tag and Twin-Strep-tag was inserted into the eukaryotic expression vector pαH (plasmid 3). Three hours after transient transfection of the plasmid into FreeStyle 293-F cells, kifunensine was added to a final concentration of 5 μM. After 6 d, supernatant was filtered and passed over a Strep-Tactin column (IBA). The column was washed with PBS, and the S1-NTD was eluted by incubating the resin with HRV3C (1% wt/wt). The S1-NTD was further purified using a Superdex 200 column (GE Healthcare). S1-NTD mutants were expressed and purified using the same method.</p><p id="p0235">Mutations were introduced into the S1-NTD fragment by overlapping PCR. To produce deglycosylated S1-NTD protein, EndoH (10% wt/wt) was added together with HRV3C (1% wt/wt) after the binding of protein onto a Strep-Tactin column.</p></sec><sec id="sec4.4.6"><title>Production of MERS-CoV S0 and S-2P</title><p id="p0240">A mammalian-codon-optimized gene encoding MERS-CoV S (England1 strain) residues 1–1291 with a C-terminal T4 fibritin trimerization domain, HRV3C cleavage site, 8xHis-tag and Twin-Strep-tag was synthesized and subcloned into the eukaryotic-expression vector pαH. The S1/S2 furin-recognition site 748-RSVR-751 was mutated to ASVG to produce a single-chain MERS-S0 protein (plasmid 4). Two proline mutations, D1059P and V1060P, were introduced to S0 to generate a stabilized MERS-CoV S-2P protein (plasmid 5). Additional mutations were introduced into MERS-CoV S-2P to make MERS-CoV S-2P variants including S-2P-S28F and S-2P-G198D.</p><p id="p0245">For expression, 0.5–1 L FreeStyle 293-F cells were transfected. Three hours after transfection, kifunensine was added to a final concentration of 5 μM. Cultures were harvested after 6 d, and protein was purified from the supernatant using Strep-Tactin resin (IBA). HRV3C protease (1% wt/wt) was added to the protein, and the reaction was incubated overnight at 4°C. Digested protein was further purified using a Superose 6 16/70 column (GE Healthcare).</p></sec><sec id="sec4.4.7"><title>Production of MERS-CoV S-NTD Bound to G2 Fab</title><p id="p0250">Different methods to produce the S1-NTD–G2 Fab complex were tested to obtain crystals, involving different S1-NTD truncations, deglycosylation strategies, co-expression and co-purification strategies. High-quality crystals were only produced by co-transfection of S1-NTD and G2 in the presence of kifunensine, yet without any glycosidase treatment, as described in more detail below.</p><p id="p0255">The three plasmids encoding G2 heavy chain (plasmid 1), G2 light chain (plasmid 2) and S1-NTD (plasmid 3) were co-transfected into FreeStyle 293-F cells. Three hours after transient transfection, kifunensine was added to a final concentration of 5 μM. After 6 d, supernatant was passed over a Protein A agarose column. The column was washed with PBS, and the complex was eluted by incubating resin with HRV3C (1% wt/wt). The sample was further purified using a Superdex 200 column (GE Healthcare).</p></sec><sec id="sec4.4.8"><title>Crystallization and X-Ray Data Collection</title><p id="p0260">Crystals of G2 Fab were produced by hanging-drop vapor diffusion by mixing 1 μL of G2 Fab (11.5 mg/mL) with 1 μL of reservoir solution containing 15% ethanol and 40% pentaerythritol propoxylate (5/4 PO/OH). Crystals were soaked in reservoir solution supplemented with 25% (v/v) glycerol and frozen in liquid nitrogen. Diffraction data were collected at the SBC beamline 19-ID (Advanced Photon Source, Argonne National Laboratory).</p><p id="p0265">Crystals of the complex of MERS-CoV S1-NTD with G2 Fab were produced by hanging-drop vapor diffusion by mixing 1 μL of protein (9.1 mg/mL) with 1 μL of reservoir solution containing 0.1 M imidazole pH 6.5, 0.2 M Li<sub>2</sub>SO<sub>4</sub>, 19% polyethylene glycol (PEG) 3350, and 6% 2-methyl-2,4-pentanediol (MPD). Crystals were soaked in reservoir solution supplemented with 20% (v/v) ethylene glycol and frozen in liquid nitrogen. Diffraction data were collected at the SBC beamline 19-ID (Advanced Photon Source, Argonne National Laboratory).</p></sec><sec id="sec4.4.9"><title>X-Ray Structure Determination and Refinement</title><p id="p0270">Diffraction data were processed using the CCP4 software suite (<xref rid="bib49" ref-type="bibr">Potterton et al., 2003</xref>): data were indexed and integrated in iMOSFLM (<xref rid="bib5" ref-type="bibr">Battye et al., 2011</xref>) and scaled and merged with AIMLESS (<xref rid="bib15" ref-type="bibr">Evans and Murshudov, 2013</xref>). A molecular replacement solution for the 2.1 Å G2 Fab dataset was found by PHASER (<xref rid="bib36" ref-type="bibr">McCoy et al., 2007</xref>) using the variable and constant domains of PDB ID: <ext-link ext-link-type="uri" xlink:href="pdb:3WBD" id="intref0130">3WBD</ext-link> (<xref rid="bib40" ref-type="bibr">Nagae et al., 2013</xref>) and PDB ID: <ext-link ext-link-type="uri" xlink:href="pdb:5VZR" id="intref0135">5VZR</ext-link> (<xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>), respectively, as search models. The structure was built manually in COOT (<xref rid="bib14" ref-type="bibr">Emsley and Cowtan, 2004</xref>) and refined using PHENIX (<xref rid="bib1" ref-type="bibr">Adams et al., 2002</xref>).</p><p id="p0275">A molecular replacement solution for the S1-NTD–G2 complex was obtained using PHASER with MERS-CoV S1-NTD (PDB ID: <ext-link ext-link-type="uri" xlink:href="pdb:5VYH" id="intref0140">5VYH</ext-link> (<xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>)) and the solved G2 Fab structure as search models. There are two S1-NTD–G2 Fab complexes in each asymmetric unit (ASU). Model building was performed in COOT and refinement was performed in PHENIX. Data collection and refinement statistics for both structures are presented in <xref rid="mmc1" ref-type="supplementary-material">Table S2</xref>.</p></sec><sec id="sec4.4.10"><title>Production of MERS-CoV S0 Bound to G2 Fab</title><p id="p0280">Three plasmids (plasmids 1, 2 and 4) were co-transfected into FreeStyle 293-F cells. Three hours after transient transfection, kifunensine was added to a final concentration of 5 μM. After 6 d, supernatant was passed over a Protein A agarose column. The column was washed with PBS and the complex was eluted by incubating resin with HRV3C (1% wt/wt). The sample was further purified using a Superose 6 16/70 column (GE Healthcare).</p></sec><sec id="sec4.4.11"><title>Negative-Stain Electron Microscopy</title><p id="p0285">Purified MERS-CoV S0–G2 Fab complex was added to carbon-coated 400 mesh copper grids and stained with 2% uranyl formate. Negative stain EM data was collected on a Tecnai Spirit operating at 120 kV with a 4k x 4k TemCam F416 camera. Micrographs were imaged through Leginon (<xref rid="bib48" ref-type="bibr">Potter et al., 1999</xref>) and processed in Appion (<xref rid="bib28" ref-type="bibr">Lander et al., 2009</xref>). Particles were selected using DoG picker (<xref rid="bib64" ref-type="bibr">Voss et al., 2009</xref>), stacked, and 2D classes were produced by MSA/MRA (<xref rid="bib43" ref-type="bibr">Ogura et al., 2003</xref>). The final 3D map was generated with EMAN2 (<xref rid="bib61" ref-type="bibr">Tang et al., 2007</xref>).</p></sec><sec id="sec4.4.12"><title>Cryo-EM Data Collection and Processing</title><p id="p0290">MERS-CoV S0–G2 complex was imaged on a Titan Krios operating at 300 kV using Leginon (<xref rid="bib48" ref-type="bibr">Potter et al., 1999</xref>, <xref rid="bib60" ref-type="bibr">Suloway et al., 2005</xref>). Each micrograph movie was collected at a magnification of 29,000x, resulting in a pixel size of 0.51 Å/pixel. Micrograph movies were recorded on a Gatan K2 detector in super-resolution mode. The dose rate was 2.50 e<sup>-</sup>/pixel/s and the defocus range was 0.5–3.5 μm. Movie frames were aligned and dose-weighted using MotionCor2 (<xref rid="bib78" ref-type="bibr">Zheng et al., 2017</xref>) and CTF models were calculated using GCTF (<xref rid="bib77" ref-type="bibr">Zhang, 2016</xref>). A small subset of particles were picked using RELION’s LoG Picker (<xref rid="bib79" ref-type="bibr">Zivanov et al., 2018</xref>), which in turn were used to prepare templates for the template picker in cryoSPARC 2 (<xref rid="bib50" ref-type="bibr">Punjani et al., 2017</xref>), resulting in 490,790 particles. 2D classification was used to clean junk particles and a clean particle stack submitted to 2-class <italic>ab initio</italic> refinement in cryoSPARC 2. The class representing the structure was used to perform one additional 2D classification step resulting in 5 templates. These templates were then used to pick particles on the original dataset resulting in 418,781 particles. The extracted particles were binned by a factor of 2. These particles were then subjected to an iterative process of filtering based on 2 class <italic>ab initio</italic> model refinement. Finally, homogeneous 3D refinement with sharpening in cryoSPARC 2 performed with a subset of 12,386 particles resulted in a 4.19 Å resolution reconstruction. Local resolution analysis performed in the latest version of cryoSPARC 2 of the map revealed a 4.44 Å resolution. Data collection and processing statistics are presented in <xref rid="mmc1" ref-type="supplementary-material">Table S3</xref>.</p></sec><sec id="sec4.4.13"><title>Cryo-EM Model Generation</title><p id="p0295">A previously published structure of MERS-CoV S (PDB ID: <ext-link ext-link-type="uri" xlink:href="pdb:5w9i" id="intref0145">5w9i</ext-link>) (<xref rid="bib44" ref-type="bibr">Pallesen et al., 2017</xref>) exhibited a good fit into our map. We then aligned the model from the NTD-G2 crystal structure to the NTD domains of PDB <ext-link ext-link-type="uri" xlink:href="pdb:5w9i" id="intref0150">5w9i</ext-link>, which also exhibited a good fit except for the constant region of the G2 Fab which is typically flexible. We therefore deleted the constant region from the model. We also deleted the NTDs from 5w9i in favor of the higher resolution NTD from the NTD-G2 crystal structure. Given the resolution of our map, in particular the lower resolution of the NTD-G2 portion, we only conducted a light refinement in Rosetta (v2019.14.60699) (<xref rid="bib9" ref-type="bibr">Conway et al., 2014</xref>) using the relax function (with constrain_relax_to_start_coords) to alleviate sidechain clashes in the hybrid model.</p></sec><sec id="sec4.4.14"><title>Surface Plasmon Resonance Affinity Assays</title><p id="p0300">A Biacore X100 (GE Healthcare) was used to measure the binding of G2 Fab to immobilized MERS-CoV S1-NTD, S1-NTD mutants, S-2P or S-2P mutants. S1-NTD or S1-NTD mutants with an 8xHis-tag and Twin-Strep-tag were immobilized on a Ni-NTA sensor chip to a total of 80–120 response units. The chip was regenerated between each cycle using 0.35 M EDTA followed by 0.5 mM NiCl<sub>2</sub>. A buffer-only sample was injected over the ligand-bound and reference flow cells, followed by G2 Fab serially diluted 2.5-fold in HBS-P<sup>+</sup> starting at 500 nM. Data were double-reference subtracted and fit to a 1:1 binding model using the Scrubber2 analysis software. To measure binding of G2 Fab to MERS-CoV S-2P or MERS-CoV S-2P mutants, similar experiments were performed, except that the S protein was immobilized to a total of 350–450 response units. All assays were performed at 25°C.</p></sec><sec id="sec4.4.15"><title>Flow Cytometry Using rDPP4-Expressing Cells</title><p id="p0305">MERS-CoV S-2P (or the S28F or G198D mutants) was fused with a C-terminal HRV3C cleavage site, a GFP tag, an 8xHis-tag and a Strep-tag II, and subcloned into the vector pαH. Proteins were expressed in FreeStyle 293-F cells and purified using Strep-Tactin column.</p><p id="p0310">FreeStyle 293-F cells were transfected with plasmids expressing full-length DPP4. 60 h after transfection, cells were harvested and washed twice with blocking buffer (PBS buffer supplemented with 0.5% BSA and 2 mM EDTA). Cells were then incubated with GFP-tagged S protein (or S variants, 200 nM) or GFP-tagged S protein (or S variants, 200 nM) supplemented with G2 or D12 or AM14 IgG (1 μM) for 30 min at 37°C. Cells were then washed twice with blocking buffer and subsequently stained with SYTOX Blue Dead Cell Stain (1:2000, ThermoFisher) before analysis using a MACSQuant VYB (Miltenyi Biotec). Data were analyzed with FlowJo software (Tree Star Inc.) using the following gating strategy: size & granularity → live cells (SYTOX Blue negative) → binding signal (GFP positive).</p></sec><sec id="sec4.4.16"><title>Flow Cytometry Using S-Expressing Cells</title><p id="p0315">Human DPP4 ectodomain (residues 39–766) was fused with an artificial signal peptide (MRPTWAWWLFLVLLLALWAPARG) at the N terminus, and a HRV3C cleavage site, 8xHis tag and Twin-Strep tag at the C terminus, followed by subcloning into the vector pαH. Protein designated as DPP4-ECD-HSS was expressed in FreeStyle 293-F cells and purified using a Strep-Tactin column.</p><p id="p0320">MERS-CoV S1 (residues 1–751) was fused with the RSV F transmembrane motif and subcloned into the vector pαH to generate the plasmid pαH-S1-TM. Full-length MERS-CoV S-WT or the stabilized S variant MERS-CoV S-2P was subcloned into the vector pαH to generate the plasmids pαH-S-WT-FL and pαH-S-2P-FL, respectively. FreeStyle 293-F cells were transfected and then harvested after 60 h to obtain S1-TM<sup>+</sup>, S-WT-FL<sup>+</sup> and S-2P-FL<sup>+</sup> cells.</p><p id="p0325">S1-TM<sup>+</sup>, S-WT-FL<sup>+</sup> and S-2P-FL<sup>+</sup> cells were then washed twice with blocking buffer (PBS buffer supplemented with 0.5% BSA and 2 mM EDTA) and subsequently incubated with 100 nM DPP4-ECD-HSS with 200 nM anti-strep antibody (produced in-house) in the presence or absence of 500 nM G2 or G4 or JC57-14 Fab for 30 min at room temperature. Cells were washed, and a 1:500 dilution of Alexa Fluor™ 647 goat anti-human IgG (H+L) (Invitrogen) was added and incubated for 30 min. Cells were then washed twice with blocking buffer and subsequently stained with SYTOX Blue Dead Cell Stain (1:2000, ThermoFisher) before analysis using a LSRFortessa SORP Flow Cytometer (BD). Data were analyzed with FlowJo software (Tree Star Inc.) using the following gating strategy: size & granularity → live cells (SYTOX Blue negative) → binding signal (Alexa Fluor™ 647 positive).</p></sec><sec id="sec4.4.17"><title>Image Cytometric Analysis</title><p id="p0330">Image cytometry methods were performed as previously described (<xref rid="bib53" ref-type="bibr">Rosen et al., 2019</xref>). Briefly, BHK-21 cells were seeded into flat bottom black-walled Greiner 96-well plates and allowed to adhere overnight. On the following day, a DPP4 expression plasmid was transfected into the cells, using Lipofectamine 3000 reagent. Two days following DPP4 transfection, MERS-CoV S-2P was incubated with 4-fold serial dilutions of antibody (Fab or IgG) for 30 min at room temperature (RT). The mixture of MERS-CoV S and antibody was then added to the DPP4-expressing BHK-21 cells and incubated for 2 hours at RT. After incubation, cells were washed, fixed with 80% cold acetone, and rewashed. Subsequently, cells were stained with MERS-CoV S rabbit polyclonal antibodies (Sino Biological, Beijing, China) and then secondary goat anti-rabbit IgG H&L labeled with Alexa Fluor® 488 (AF488) was added. Finally, cell nuclei were stained with DAPI. Percent inhibition as a function of antibody concentrations was then plotted and analyzed via a one-site-fit Log IC<sub>50</sub> non-linear regression analysis. No inhibition (0%) was defined as MERS-CoV S binding to BHK-21 cells without the addition of antibody. Full inhibition (100%) was defined as MERS-CoV S binding to BHK-21 without DPP4 receptor.</p></sec><sec id="sec4.4.18"><title>Neutralization Assay Comparing G2 IgG versus Fab</title><p id="p0335">MERS-CoV (England1 strain) pseudovirions, expressing wild-type and mutant S proteins, were produced by co-transfection of three plasmids into 293T cells using Fugene 6 transfection reagent (Promega, Madison, WI) and titered, as described previously (<xref rid="bib69" ref-type="bibr">Wang et al., 2018</xref>). For the neutralization assay, BHK-21 cells with exogenously expressed DPP4 were used as previously described (<xref rid="bib53" ref-type="bibr">Rosen et al., 2019</xref>). Briefly, DPP4 was expressed in BHK-21 cells and two days post-transfection, antibodies were mixed with MERS-CoV pseudoviruses and added to cells. 72 hours later, cells were lysed and analyzed for luciferase activity. Relative luciferase units were measured and percent neutralization was calculated considering uninfected cells as 100% neutralization and cells transduced with only pseudovirus as 0% neutralization.</p></sec><sec id="sec4.4.19"><title>Surface Plasmon Resonance Competition Assay</title><p id="p0340">A Biacore X100 (GE Healthcare) was used to measure the binding of un-tagged MERS-CoV S-2P—alone or in the presence of a 5-fold molar excess of IgG or Fab—to immobilized twin-Strep-tagged DPP4 ectodomain (DPP4-ECD-HSS). DPP4-ECD-HSS was captured using an anti-Strep antibody-coupled CM5 chip to a total of 65–75 response units. The chip was regenerated between each cycle using 10 mM glycine, pH 2.0 followed by 0.1% SDS. HBS-P<sup>+</sup> pH 8.0 was used for running buffer as well as sample buffer. For each cycle, HBS P<sup>+</sup> pH 8.0 was injected once after immobilization to further clean the chip, followed by the injection of buffer only, 500 nM Fab or IgG only, 100 nM S-2P only, or 100 nM S-2P supplemented with 500 nM Fab or IgG. Experiments were performed at 25°C. Data were reference-subtracted and analyzed in BIAevaluation software.</p></sec></sec><sec id="sec4.5"><title>Quantification and Statistical Analysis</title><p id="p0345">Binding and neutralization assays were conducted with at least duplicate measurements and presented as the mean ± SEM of the indicated number of replicates. Details can be found in figure legends.</p></sec><sec sec-type="data-availability" id="sec4.6"><title>Data and Code Availability</title><p id="p0350">Coordinates and structure factors for G2 Fab have been deposited in the Protein Data Bank under accession code PDB: <ext-link ext-link-type="uri" xlink:href="pdb:6PXG" id="intref0155">6PXG</ext-link>. Coordinates and structure factors for MERS-CoV S1-NTD–G2 Fab have been deposited in the Protein Data Bank under accession code PDB: <ext-link ext-link-type="uri" xlink:href="pdb:6PXH" id="intref0160">6PXH</ext-link>. Cryo-EM reconstruction of MERS-S0–G2 Fab complex has been deposited in the Electron Microscopy Data Bank (EMDB) (accession code EMDB: EMD-20527). Atomic models have been deposited in the Protein Data Bank (PDB: <ext-link ext-link-type="uri" xlink:href="pdb:6PZ8" id="intref0165">6PZ8</ext-link>).</p></sec></sec></body><back><ref-list id="cebib0010"><title>References</title><ref id="bib1"><element-citation publication-type="journal" id="sref1"><person-group person-group-type="author"><name><surname>Adams</surname><given-names>P.D.</given-names></name><name><surname>Grosse-Kunstleve</surname><given-names>R.W.</given-names></name><name><surname>Hung</surname><given-names>L.W.</given-names></name><name><surname>Ioerger</surname><given-names>T.R.</given-names></name><name><surname>McCoy</surname><given-names>A.J.</given-names></name><name><surname>Moriarty</surname><given-names>N.W.</given-names></name><name><surname>Read</surname><given-names>R.J.</given-names></name><name><surname>Sacchettini</surname><given-names>J.C.</given-names></name><name><surname>Sauter</surname><given-names>N.K.</given-names></name><name><surname>Terwilliger</surname><given-names>T.C.</given-names></name></person-group><article-title>PHENIX: building new software for automated crystallographic structure determination</article-title><source>Acta. Crystallogr. D Biol. Crystallogr.</source><volume>58</volume><year>2002</year><fpage>1948</fpage><lpage>1954</lpage><pub-id pub-id-type="pmid">12393927</pub-id></element-citation></ref><ref id="bib2"><element-citation publication-type="journal" id="sref2"><person-group person-group-type="author"><name><surname>Almazán</surname><given-names>F.</given-names></name><name><surname>DeDiego</surname><given-names>M.L.</given-names></name><name><surname>Sola</surname><given-names>I.</given-names></name><name><surname>Zuñiga</surname><given-names>S.</given-names></name><name><surname>Nieto-Torres</surname><given-names>J.L.</given-names></name><name><surname>Marquez-Jurado</surname><given-names>S.</given-names></name><name><surname>Andrés</surname><given-names>G.</given-names></name><name><surname>Enjuanes</surname><given-names>L.</given-names></name></person-group><article-title>Engineering a replication-competent, propagation-defective Middle East respiratory syndrome coronavirus as a vaccine candidate</article-title><source>MBio</source><volume>4</volume><year>2013</year><fpage>e00650-13</fpage><pub-id pub-id-type="pmid">24023385</pub-id></element-citation></ref><ref id="bib3"><element-citation publication-type="journal" id="sref3"><person-group person-group-type="author"><name><surname>Assiri</surname><given-names>A.M.</given-names></name><name><surname>Midgley</surname><given-names>C.M.</given-names></name><name><surname>Abedi</surname><given-names>G.R.</given-names></name><name><surname>Bin Saeed</surname><given-names>A.</given-names></name><name><surname>Almasri</surname><given-names>M.M.</given-names></name><name><surname>Lu</surname><given-names>X.</given-names></name><name><surname>Al-Abdely</surname><given-names>H.M.</given-names></name><name><surname>Abdalla</surname><given-names>O.</given-names></name><name><surname>Mohammed</surname><given-names>M.</given-names></name><name><surname>Algarni</surname><given-names>H.S.</given-names></name></person-group><article-title>Epidemiology of a Novel Recombinant Middle East Respiratory Syndrome Coronavirus in Humans in Saudi Arabia</article-title><source>J. Infect. Dis.</source><volume>214</volume><year>2016</year><fpage>712</fpage><lpage>721</lpage><pub-id pub-id-type="pmid">27302191</pub-id></element-citation></ref><ref id="bib4"><element-citation publication-type="journal" id="sref4"><person-group person-group-type="author"><name><surname>Azhar</surname><given-names>E.I.</given-names></name><name><surname>El-Kafrawy</surname><given-names>S.A.</given-names></name><name><surname>Farraj</surname><given-names>S.A.</given-names></name><name><surname>Hassan</surname><given-names>A.M.</given-names></name><name><surname>Al-Saeed</surname><given-names>M.S.</given-names></name><name><surname>Hashem</surname><given-names>A.M.</given-names></name><name><surname>Madani</surname><given-names>T.A.</given-names></name></person-group><article-title>Evidence for camel-to-human transmission of MERS coronavirus</article-title><source>N. Engl. J. Med.</source><volume>370</volume><year>2014</year><fpage>2499</fpage><lpage>2505</lpage><pub-id pub-id-type="pmid">24896817</pub-id></element-citation></ref><ref id="bib5"><element-citation publication-type="journal" id="sref5"><person-group person-group-type="author"><name><surname>Battye</surname><given-names>T.G.</given-names></name><name><surname>Kontogiannis</surname><given-names>L.</given-names></name><name><surname>Johnson</surname><given-names>O.</given-names></name><name><surname>Powell</surname><given-names>H.R.</given-names></name><name><surname>Leslie</surname><given-names>A.G.</given-names></name></person-group><article-title>iMOSFLM: a new graphical interface for diffraction-image processing with MOSFLM</article-title><source>Acta Crystallogr. D Biol. Crystallogr.</source><volume>67</volume><year>2011</year><fpage>271</fpage><lpage>281</lpage><pub-id pub-id-type="pmid">21460445</pub-id></element-citation></ref><ref id="bib6"><element-citation publication-type="journal" id="sref6"><person-group person-group-type="author"><name><surname>Chan</surname><given-names>C.M.</given-names></name><name><surname>Chu</surname><given-names>H.</given-names></name><name><surname>Wang</surname><given-names>Y.</given-names></name><name><surname>Wong</surname><given-names>B.H.</given-names></name><name><surname>Zhao</surname><given-names>X.</given-names></name><name><surname>Zhou</surname><given-names>J.</given-names></name><name><surname>Yang</surname><given-names>D.</given-names></name><name><surname>Leung</surname><given-names>S.P.</given-names></name><name><surname>Chan</surname><given-names>J.F.</given-names></name><name><surname>Yeung</surname><given-names>M.L.</given-names></name></person-group><article-title>Carcinoembryonic Antigen-Related Cell Adhesion Molecule 5 Is an Important Surface Attachment Factor That Facilitates Entry of Middle East Respiratory Syndrome Coronavirus</article-title><source>J. Virol.</source><volume>90</volume><year>2016</year><fpage>9114</fpage><lpage>9127</lpage><pub-id pub-id-type="pmid">27489282</pub-id></element-citation></ref><ref id="bib7"><element-citation publication-type="journal" id="sref7"><person-group person-group-type="author"><name><surname>Chen</surname><given-names>Y.</given-names></name><name><surname>Lu</surname><given-names>S.</given-names></name><name><surname>Jia</surname><given-names>H.</given-names></name><name><surname>Deng</surname><given-names>Y.</given-names></name><name><surname>Zhou</surname><given-names>J.</given-names></name><name><surname>Huang</surname><given-names>B.</given-names></name><name><surname>Yu</surname><given-names>Y.</given-names></name><name><surname>Lan</surname><given-names>J.</given-names></name><name><surname>Wang</surname><given-names>W.</given-names></name><name><surname>Lou</surname><given-names>Y.</given-names></name></person-group><article-title>A novel neutralizing monoclonal antibody targeting the N-terminal domain of the MERS-CoV spike protein</article-title><source>Emerg. Microbes Infect.</source><volume>6</volume><year>2017</year><fpage>e60</fpage><pub-id pub-id-type="pmid">28655936</pub-id></element-citation></ref><ref id="bib8"><element-citation publication-type="journal" id="sref8"><person-group person-group-type="author"><name><surname>Chu</surname><given-names>H.</given-names></name><name><surname>Chan</surname><given-names>C.M.</given-names></name><name><surname>Zhang</surname><given-names>X.</given-names></name><name><surname>Wang</surname><given-names>Y.</given-names></name><name><surname>Yuan</surname><given-names>S.</given-names></name><name><surname>Zhou</surname><given-names>J.</given-names></name><name><surname>Au-Yeung</surname><given-names>R.K.</given-names></name><name><surname>Sze</surname><given-names>K.H.</given-names></name><name><surname>Yang</surname><given-names>D.</given-names></name><name><surname>Shuai</surname><given-names>H.</given-names></name></person-group><article-title>Middle East respiratory syndrome coronavirus and bat coronavirus HKU9 both can utilize GRP78 for attachment onto host cells</article-title><source>J. Biol. Chem.</source><volume>293</volume><year>2018</year><fpage>11709</fpage><lpage>11726</lpage><pub-id pub-id-type="pmid">29887526</pub-id></element-citation></ref><ref id="bib9"><element-citation publication-type="journal" id="sref9"><person-group person-group-type="author"><name><surname>Conway</surname><given-names>P.</given-names></name><name><surname>Tyka</surname><given-names>M.D.</given-names></name><name><surname>DiMaio</surname><given-names>F.</given-names></name><name><surname>Konerding</surname><given-names>D.E.</given-names></name><name><surname>Baker</surname><given-names>D.</given-names></name></person-group><article-title>Relaxation of backbone bond geometry improves protein energy landscape modeling</article-title><source>Protein Sci.</source><volume>23</volume><year>2014</year><fpage>47</fpage><lpage>55</lpage><pub-id pub-id-type="pmid">24265211</pub-id></element-citation></ref><ref id="bib10"><element-citation publication-type="journal" id="sref10"><person-group person-group-type="author"><name><surname>Corti</surname><given-names>D.</given-names></name><name><surname>Zhao</surname><given-names>J.</given-names></name><name><surname>Pedotti</surname><given-names>M.</given-names></name><name><surname>Simonelli</surname><given-names>L.</given-names></name><name><surname>Agnihothram</surname><given-names>S.</given-names></name><name><surname>Fett</surname><given-names>C.</given-names></name><name><surname>Fernandez-Rodriguez</surname><given-names>B.</given-names></name><name><surname>Foglierini</surname><given-names>M.</given-names></name><name><surname>Agatic</surname><given-names>G.</given-names></name><name><surname>Vanzetta</surname><given-names>F.</given-names></name></person-group><article-title>Prophylactic and postexposure efficacy of a potent human monoclonal antibody against MERS coronavirus</article-title><source>Proc. Natl. Acad. Sci. USA</source><volume>112</volume><year>2015</year><fpage>10473</fpage><lpage>10478</lpage><pub-id pub-id-type="pmid">26216974</pub-id></element-citation></ref><ref id="bib11"><element-citation publication-type="journal" id="sref11"><person-group person-group-type="author"><name><surname>Coughlin</surname><given-names>M.M.</given-names></name><name><surname>Babcook</surname><given-names>J.</given-names></name><name><surname>Prabhakar</surname><given-names>B.S.</given-names></name></person-group><article-title>Human monoclonal antibodies to SARS-coronavirus inhibit infection by different mechanisms</article-title><source>Virology</source><volume>394</volume><year>2009</year><fpage>39</fpage><lpage>46</lpage><pub-id pub-id-type="pmid">19748648</pub-id></element-citation></ref><ref id="bib12"><element-citation publication-type="journal" id="sref12"><person-group person-group-type="author"><name><surname>Drosten</surname><given-names>C.</given-names></name><name><surname>Muth</surname><given-names>D.</given-names></name><name><surname>Corman</surname><given-names>V.M.</given-names></name><name><surname>Hussain</surname><given-names>R.</given-names></name><name><surname>Al Masri</surname><given-names>M.</given-names></name><name><surname>HajOmar</surname><given-names>W.</given-names></name><name><surname>Landt</surname><given-names>O.</given-names></name><name><surname>Assiri</surname><given-names>A.</given-names></name><name><surname>Eckerle</surname><given-names>I.</given-names></name><name><surname>Al Shangiti</surname><given-names>A.</given-names></name></person-group><article-title>An observational, laboratory-based study of outbreaks of middle East respiratory syndrome coronavirus in Jeddah and Riyadh, kingdom of Saudi Arabia, 2014</article-title><source>Clin. Infect. Dis.</source><volume>60</volume><year>2015</year><fpage>369</fpage><lpage>377</lpage><pub-id pub-id-type="pmid">25323704</pub-id></element-citation></ref><ref id="bib13"><element-citation publication-type="journal" id="sref13"><person-group person-group-type="author"><name><surname>Du</surname><given-names>L.</given-names></name><name><surname>Tai</surname><given-names>W.</given-names></name><name><surname>Yang</surname><given-names>Y.</given-names></name><name><surname>Zhao</surname><given-names>G.</given-names></name><name><surname>Zhu</surname><given-names>Q.</given-names></name><name><surname>Sun</surname><given-names>S.</given-names></name><name><surname>Liu</surname><given-names>C.</given-names></name><name><surname>Tao</surname><given-names>X.</given-names></name><name><surname>Tseng</surname><given-names>C.K.</given-names></name><name><surname>Perlman</surname><given-names>S.</given-names></name></person-group><article-title>Introduction of neutralizing immunogenicity index to the rational design of MERS coronavirus subunit vaccines</article-title><source>Nat. Commun.</source><volume>7</volume><year>2016</year><fpage>13473</fpage><pub-id pub-id-type="pmid">27874853</pub-id></element-citation></ref><ref id="bib14"><element-citation publication-type="journal" id="sref14"><person-group person-group-type="author"><name><surname>Emsley</surname><given-names>P.</given-names></name><name><surname>Cowtan</surname><given-names>K.</given-names></name></person-group><article-title>Coot: model-building tools for molecular graphics</article-title><source>Acta Crystallogr. D Biol. Crystallogr.</source><volume>60</volume><year>2004</year><fpage>2126</fpage><lpage>2132</lpage><pub-id pub-id-type="pmid">15572765</pub-id></element-citation></ref><ref id="bib15"><element-citation publication-type="journal" id="sref15"><person-group person-group-type="author"><name><surname>Evans</surname><given-names>P.R.</given-names></name><name><surname>Murshudov</surname><given-names>G.N.</given-names></name></person-group><article-title>How good are my data and what is the resolution?</article-title><source>Acta Crystallogr. D Biol. Crystallogr.</source><volume>69</volume><year>2013</year><fpage>1204</fpage><lpage>1214</lpage><pub-id pub-id-type="pmid">23793146</pub-id></element-citation></ref><ref id="bib16"><element-citation publication-type="journal" id="sref16"><person-group person-group-type="author"><name><surname>Fehr</surname><given-names>A.R.</given-names></name><name><surname>Athmer</surname><given-names>J.</given-names></name><name><surname>Channappanavar</surname><given-names>R.</given-names></name><name><surname>Phillips</surname><given-names>J.M.</given-names></name><name><surname>Meyerholz</surname><given-names>D.K.</given-names></name><name><surname>Perlman</surname><given-names>S.</given-names></name></person-group><article-title>The nsp3 macrodomain promotes virulence in mice with coronavirus-induced encephalitis</article-title><source>J. Virol.</source><volume>89</volume><year>2015</year><fpage>1523</fpage><lpage>1536</lpage><pub-id pub-id-type="pmid">25428866</pub-id></element-citation></ref><ref id="bib17"><element-citation publication-type="journal" id="sref17"><person-group person-group-type="author"><name><surname>Gierer</surname><given-names>S.</given-names></name><name><surname>Bertram</surname><given-names>S.</given-names></name><name><surname>Kaup</surname><given-names>F.</given-names></name><name><surname>Wrensch</surname><given-names>F.</given-names></name><name><surname>Heurich</surname><given-names>A.</given-names></name><name><surname>Krämer-Kühl</surname><given-names>A.</given-names></name><name><surname>Welsch</surname><given-names>K.</given-names></name><name><surname>Winkler</surname><given-names>M.</given-names></name><name><surname>Meyer</surname><given-names>B.</given-names></name><name><surname>Drosten</surname><given-names>C.</given-names></name></person-group><article-title>The spike protein of the emerging betacoronavirus EMC uses a novel coronavirus receptor for entry, can be activated by TMPRSS2, and is targeted by neutralizing antibodies</article-title><source>J. Virol.</source><volume>87</volume><year>2013</year><fpage>5502</fpage><lpage>5511</lpage><pub-id pub-id-type="pmid">23468491</pub-id></element-citation></ref><ref id="bib18"><element-citation publication-type="journal" id="sref18"><person-group person-group-type="author"><name><surname>Greenough</surname><given-names>T.C.</given-names></name><name><surname>Babcock</surname><given-names>G.J.</given-names></name><name><surname>Roberts</surname><given-names>A.</given-names></name><name><surname>Hernandez</surname><given-names>H.J.</given-names></name><name><surname>Thomas</surname><given-names>W.D.</given-names><suffix>Jr.</suffix></name><name><surname>Coccia</surname><given-names>J.A.</given-names></name><name><surname>Graziano</surname><given-names>R.F.</given-names></name><name><surname>Srinivasan</surname><given-names>M.</given-names></name><name><surname>Lowy</surname><given-names>I.</given-names></name><name><surname>Finberg</surname><given-names>R.W.</given-names></name></person-group><article-title>Development and characterization of a severe acute respiratory syndrome-associated coronavirus-neutralizing human monoclonal antibody that provides effective immunoprophylaxis in mice</article-title><source>J. Infect. Dis.</source><volume>191</volume><year>2005</year><fpage>507</fpage><lpage>514</lpage><pub-id pub-id-type="pmid">15655773</pub-id></element-citation></ref><ref id="bib19"><element-citation publication-type="journal" id="sref19"><person-group person-group-type="author"><name><surname>Gui</surname><given-names>M.</given-names></name><name><surname>Song</surname><given-names>W.</given-names></name><name><surname>Zhou</surname><given-names>H.</given-names></name><name><surname>Xu</surname><given-names>J.</given-names></name><name><surname>Chen</surname><given-names>S.</given-names></name><name><surname>Xiang</surname><given-names>Y.</given-names></name><name><surname>Wang</surname><given-names>X.</given-names></name></person-group><article-title>Cryo-electron microscopy structures of the SARS-CoV spike glycoprotein reveal a prerequisite conformational state for receptor binding</article-title><source>Cell Res.</source><volume>27</volume><year>2017</year><fpage>119</fpage><lpage>129</lpage><pub-id pub-id-type="pmid">28008928</pub-id></element-citation></ref><ref id="bib20"><element-citation publication-type="journal" id="sref20"><person-group person-group-type="author"><name><surname>Guo</surname><given-names>Y.</given-names></name><name><surname>Tisoncik</surname><given-names>J.</given-names></name><name><surname>McReynolds</surname><given-names>S.</given-names></name><name><surname>Farzan</surname><given-names>M.</given-names></name><name><surname>Prabhakar</surname><given-names>B.S.</given-names></name><name><surname>Gallagher</surname><given-names>T.</given-names></name><name><surname>Rong</surname><given-names>L.</given-names></name><name><surname>Caffrey</surname><given-names>M.</given-names></name></person-group><article-title>Identification of a new region of SARS-CoV S protein critical for viral entry</article-title><source>J. Mol. Biol.</source><volume>394</volume><year>2009</year><fpage>600</fpage><lpage>605</lpage><pub-id pub-id-type="pmid">19853613</pub-id></element-citation></ref><ref id="bib21"><element-citation publication-type="journal" id="sref21"><person-group person-group-type="author"><name><surname>Jiaming</surname><given-names>L.</given-names></name><name><surname>Yanfeng</surname><given-names>Y.</given-names></name><name><surname>Yao</surname><given-names>D.</given-names></name><name><surname>Yawei</surname><given-names>H.</given-names></name><name><surname>Linlin</surname><given-names>B.</given-names></name><name><surname>Baoying</surname><given-names>H.</given-names></name><name><surname>Jinghua</surname><given-names>Y.</given-names></name><name><surname>Gao</surname><given-names>G.F.</given-names></name><name><surname>Chuan</surname><given-names>Q.</given-names></name><name><surname>Wenjie</surname><given-names>T.</given-names></name></person-group><article-title>The recombinant N-terminal domain of spike proteins is a potential vaccine against Middle East respiratory syndrome coronavirus (MERS-CoV) infection</article-title><source>Vaccine</source><volume>35</volume><year>2017</year><fpage>10</fpage><lpage>18</lpage><pub-id pub-id-type="pmid">27899228</pub-id></element-citation></ref><ref id="bib22"><element-citation publication-type="journal" id="sref22"><person-group person-group-type="author"><name><surname>Jiang</surname><given-names>L.</given-names></name><name><surname>Wang</surname><given-names>N.</given-names></name><name><surname>Zuo</surname><given-names>T.</given-names></name><name><surname>Shi</surname><given-names>X.</given-names></name><name><surname>Poon</surname><given-names>K.M.</given-names></name><name><surname>Wu</surname><given-names>Y.</given-names></name><name><surname>Gao</surname><given-names>F.</given-names></name><name><surname>Li</surname><given-names>D.</given-names></name><name><surname>Wang</surname><given-names>R.</given-names></name><name><surname>Guo</surname><given-names>J.</given-names></name></person-group><article-title>Potent neutralization of MERS-CoV by human neutralizing monoclonal antibodies to the viral spike glycoprotein</article-title><source>Sci. Transl. Med.</source><volume>6</volume><year>2014</year><fpage>234ra59</fpage></element-citation></ref><ref id="bib23"><element-citation publication-type="journal" id="sref23"><person-group person-group-type="author"><name><surname>Ki</surname><given-names>M.</given-names></name></person-group><article-title>2015 MERS outbreak in Korea: hospital-to-hospital transmission</article-title><source>Epidemiol. Health</source><volume>37</volume><year>2015</year><fpage>e2015033</fpage><pub-id pub-id-type="pmid">26212508</pub-id></element-citation></ref><ref id="bib24"><element-citation publication-type="journal" id="sref24"><person-group person-group-type="author"><name><surname>Kirchdoerfer</surname><given-names>R.N.</given-names></name><name><surname>Cottrell</surname><given-names>C.A.</given-names></name><name><surname>Wang</surname><given-names>N.</given-names></name><name><surname>Pallesen</surname><given-names>J.</given-names></name><name><surname>Yassine</surname><given-names>H.M.</given-names></name><name><surname>Turner</surname><given-names>H.L.</given-names></name><name><surname>Corbett</surname><given-names>K.S.</given-names></name><name><surname>Graham</surname><given-names>B.S.</given-names></name><name><surname>McLellan</surname><given-names>J.S.</given-names></name><name><surname>Ward</surname><given-names>A.B.</given-names></name></person-group><article-title>Pre-fusion structure of a human coronavirus spike protein</article-title><source>Nature</source><volume>531</volume><year>2016</year><fpage>118</fpage><lpage>121</lpage><pub-id pub-id-type="pmid">26935699</pub-id></element-citation></ref><ref id="bib25"><element-citation publication-type="journal" id="sref25"><person-group person-group-type="author"><name><surname>Krempl</surname><given-names>C.</given-names></name><name><surname>Schultze</surname><given-names>B.</given-names></name><name><surname>Laude</surname><given-names>H.</given-names></name><name><surname>Herrler</surname><given-names>G.</given-names></name></person-group><article-title>Point mutations in the S protein connect the sialic acid binding activity with the enteropathogenicity of transmissible gastroenteritis coronavirus</article-title><source>J. Virol.</source><volume>71</volume><year>1997</year><fpage>3285</fpage><lpage>3287</lpage><pub-id pub-id-type="pmid">9060696</pub-id></element-citation></ref><ref id="bib26"><element-citation publication-type="journal" id="sref26"><person-group person-group-type="author"><name><surname>Krissinel</surname><given-names>E.</given-names></name><name><surname>Henrick</surname><given-names>K.</given-names></name></person-group><article-title>Inference of macromolecular assemblies from crystalline state</article-title><source>J. Mol. Biol.</source><volume>372</volume><year>2007</year><fpage>774</fpage><lpage>797</lpage><pub-id pub-id-type="pmid">17681537</pub-id></element-citation></ref><ref id="bib27"><element-citation publication-type="journal" id="sref27"><person-group person-group-type="author"><name><surname>Künkel</surname><given-names>F.</given-names></name><name><surname>Herrler</surname><given-names>G.</given-names></name></person-group><article-title>Structural and functional analysis of the surface protein of human coronavirus OC43</article-title><source>Virology</source><volume>195</volume><year>1993</year><fpage>195</fpage><lpage>202</lpage><pub-id pub-id-type="pmid">8317096</pub-id></element-citation></ref><ref id="bib28"><element-citation publication-type="journal" id="sref28"><person-group person-group-type="author"><name><surname>Lander</surname><given-names>G.C.</given-names></name><name><surname>Stagg</surname><given-names>S.M.</given-names></name><name><surname>Voss</surname><given-names>N.R.</given-names></name><name><surname>Cheng</surname><given-names>A.</given-names></name><name><surname>Fellmann</surname><given-names>D.</given-names></name><name><surname>Pulokas</surname><given-names>J.</given-names></name><name><surname>Yoshioka</surname><given-names>C.</given-names></name><name><surname>Irving</surname><given-names>C.</given-names></name><name><surname>Mulder</surname><given-names>A.</given-names></name><name><surname>Lau</surname><given-names>P.W.</given-names></name></person-group><article-title>Appion: an integrated, database-driven pipeline to facilitate EM image processing</article-title><source>J. Struct. Biol.</source><volume>166</volume><year>2009</year><fpage>95</fpage><lpage>102</lpage><pub-id pub-id-type="pmid">19263523</pub-id></element-citation></ref><ref id="bib29"><element-citation publication-type="journal" id="sref29"><person-group person-group-type="author"><name><surname>Li</surname><given-names>F.</given-names></name></person-group><article-title>Receptor recognition mechanisms of coronaviruses: a decade of structural studies</article-title><source>J. Virol.</source><volume>89</volume><year>2015</year><fpage>1954</fpage><lpage>1964</lpage><pub-id pub-id-type="pmid">25428871</pub-id></element-citation></ref><ref id="bib30"><element-citation publication-type="journal" id="sref30"><person-group person-group-type="author"><name><surname>Li</surname><given-names>F.</given-names></name></person-group><article-title>Structure, Function, and Evolution of Coronavirus Spike Proteins</article-title><source>Annu. Rev. Virol.</source><volume>3</volume><year>2016</year><fpage>237</fpage><lpage>261</lpage><pub-id pub-id-type="pmid">27578435</pub-id></element-citation></ref><ref id="bib31"><element-citation publication-type="journal" id="sref31"><person-group person-group-type="author"><name><surname>Li</surname><given-names>F.</given-names></name><name><surname>Li</surname><given-names>W.</given-names></name><name><surname>Farzan</surname><given-names>M.</given-names></name><name><surname>Harrison</surname><given-names>S.C.</given-names></name></person-group><article-title>Structure of SARS coronavirus spike receptor-binding domain complexed with receptor</article-title><source>Science</source><volume>309</volume><year>2005</year><fpage>1864</fpage><lpage>1868</lpage><pub-id pub-id-type="pmid">16166518</pub-id></element-citation></ref><ref id="bib32"><element-citation publication-type="journal" id="sref32"><person-group person-group-type="author"><name><surname>Li</surname><given-names>W.</given-names></name><name><surname>Moore</surname><given-names>M.J.</given-names></name><name><surname>Vasilieva</surname><given-names>N.</given-names></name><name><surname>Sui</surname><given-names>J.</given-names></name><name><surname>Wong</surname><given-names>S.K.</given-names></name><name><surname>Berne</surname><given-names>M.A.</given-names></name><name><surname>Somasundaran</surname><given-names>M.</given-names></name><name><surname>Sullivan</surname><given-names>J.L.</given-names></name><name><surname>Luzuriaga</surname><given-names>K.</given-names></name><name><surname>Greenough</surname><given-names>T.C.</given-names></name></person-group><article-title>Angiotensin-converting enzyme 2 is a functional receptor for the SARS coronavirus</article-title><source>Nature</source><volume>426</volume><year>2003</year><fpage>450</fpage><lpage>454</lpage><pub-id pub-id-type="pmid">14647384</pub-id></element-citation></ref><ref id="bib33"><element-citation publication-type="journal" id="sref33"><person-group person-group-type="author"><name><surname>Li</surname><given-names>W.</given-names></name><name><surname>Hulswit</surname><given-names>R.J.G.</given-names></name><name><surname>Widjaja</surname><given-names>I.</given-names></name><name><surname>Raj</surname><given-names>V.S.</given-names></name><name><surname>McBride</surname><given-names>R.</given-names></name><name><surname>Peng</surname><given-names>W.</given-names></name><name><surname>Widagdo</surname><given-names>W.</given-names></name><name><surname>Tortorici</surname><given-names>M.A.</given-names></name><name><surname>van Dieren</surname><given-names>B.</given-names></name><name><surname>Lang</surname><given-names>Y.</given-names></name></person-group><article-title>Identification of sialic acid-binding function for the Middle East respiratory syndrome coronavirus spike glycoprotein</article-title><source>Proc. Natl. Acad. Sci. USA</source><volume>114</volume><year>2017</year><fpage>E8508</fpage><lpage>E8517</lpage><pub-id pub-id-type="pmid">28923942</pub-id></element-citation></ref><ref id="bib34"><element-citation publication-type="journal" id="sref34"><person-group person-group-type="author"><name><surname>Li</surname><given-names>Y.</given-names></name><name><surname>Wan</surname><given-names>Y.</given-names></name><name><surname>Liu</surname><given-names>P.</given-names></name><name><surname>Zhao</surname><given-names>J.</given-names></name><name><surname>Lu</surname><given-names>G.</given-names></name><name><surname>Qi</surname><given-names>J.</given-names></name><name><surname>Wang</surname><given-names>Q.</given-names></name><name><surname>Lu</surname><given-names>X.</given-names></name><name><surname>Wu</surname><given-names>Y.</given-names></name><name><surname>Liu</surname><given-names>W.</given-names></name></person-group><article-title>A humanized neutralizing antibody against MERS-CoV targeting the receptor-binding domain of the spike protein</article-title><source>Cell Res.</source><volume>25</volume><year>2015</year><fpage>1237</fpage><lpage>1249</lpage><pub-id pub-id-type="pmid">26391698</pub-id></element-citation></ref><ref id="bib35"><element-citation publication-type="journal" id="sref35"><person-group person-group-type="author"><name><surname>Lu</surname><given-names>G.</given-names></name><name><surname>Hu</surname><given-names>Y.</given-names></name><name><surname>Wang</surname><given-names>Q.</given-names></name><name><surname>Qi</surname><given-names>J.</given-names></name><name><surname>Gao</surname><given-names>F.</given-names></name><name><surname>Li</surname><given-names>Y.</given-names></name><name><surname>Zhang</surname><given-names>Y.</given-names></name><name><surname>Zhang</surname><given-names>W.</given-names></name><name><surname>Yuan</surname><given-names>Y.</given-names></name><name><surname>Bao</surname><given-names>J.</given-names></name></person-group><article-title>Molecular basis of binding between novel human coronavirus MERS-CoV and its receptor CD26</article-title><source>Nature</source><volume>500</volume><year>2013</year><fpage>227</fpage><lpage>231</lpage><pub-id pub-id-type="pmid">23831647</pub-id></element-citation></ref><ref id="bib36"><element-citation publication-type="journal" id="sref36"><person-group person-group-type="author"><name><surname>McCoy</surname><given-names>A.J.</given-names></name><name><surname>Grosse-Kunstleve</surname><given-names>R.W.</given-names></name><name><surname>Adams</surname><given-names>P.D.</given-names></name><name><surname>Winn</surname><given-names>M.D.</given-names></name><name><surname>Storoni</surname><given-names>L.C.</given-names></name><name><surname>Read</surname><given-names>R.J.</given-names></name></person-group><article-title>Phaser crystallographic software</article-title><source>J. Appl. Cryst.</source><volume>40</volume><year>2007</year><fpage>658</fpage><lpage>674</lpage><pub-id pub-id-type="pmid">19461840</pub-id></element-citation></ref><ref id="bib37"><element-citation publication-type="journal" id="sref37"><person-group person-group-type="author"><name><surname>Millet</surname><given-names>J.K.</given-names></name><name><surname>Whittaker</surname><given-names>G.R.</given-names></name></person-group><article-title>Host cell entry of Middle East respiratory syndrome coronavirus after two-step, furin-mediated activation of the spike protein</article-title><source>Proc. Natl. Acad. Sci. USA</source><volume>111</volume><year>2014</year><fpage>15214</fpage><lpage>15219</lpage><pub-id pub-id-type="pmid">25288733</pub-id></element-citation></ref><ref id="bib38"><element-citation publication-type="journal" id="sref38"><person-group person-group-type="author"><name><surname>Modjarrad</surname><given-names>K.</given-names></name><name><surname>Moorthy</surname><given-names>V.S.</given-names></name><name><surname>Ben Embarek</surname><given-names>P.</given-names></name><name><surname>Van Kerkhove</surname><given-names>M.</given-names></name><name><surname>Kim</surname><given-names>J.</given-names></name><name><surname>Kieny</surname><given-names>M.P.</given-names></name></person-group><article-title>A roadmap for MERS-CoV research and product development: report from a World Health Organization consultation</article-title><source>Nat. Med.</source><volume>22</volume><year>2016</year><fpage>701</fpage><lpage>705</lpage><pub-id pub-id-type="pmid">27387881</pub-id></element-citation></ref><ref id="bib39"><element-citation publication-type="journal" id="sref39"><person-group person-group-type="author"><name><surname>Mohd</surname><given-names>H.A.</given-names></name><name><surname>Al-Tawfiq</surname><given-names>J.A.</given-names></name><name><surname>Memish</surname><given-names>Z.A.</given-names></name></person-group><article-title>Middle East Respiratory Syndrome Coronavirus (MERS-CoV) origin and animal reservoir</article-title><source>Virol. J.</source><volume>13</volume><year>2016</year><fpage>87</fpage><pub-id pub-id-type="pmid">27255185</pub-id></element-citation></ref><ref id="bib40"><element-citation publication-type="journal" id="sref40"><person-group person-group-type="author"><name><surname>Nagae</surname><given-names>M.</given-names></name><name><surname>Ikeda</surname><given-names>A.</given-names></name><name><surname>Hane</surname><given-names>M.</given-names></name><name><surname>Hanashima</surname><given-names>S.</given-names></name><name><surname>Kitajima</surname><given-names>K.</given-names></name><name><surname>Sato</surname><given-names>C.</given-names></name><name><surname>Yamaguchi</surname><given-names>Y.</given-names></name></person-group><article-title>Crystal structure of anti-polysialic acid antibody single chain Fv fragment complexed with octasialic acid: insight into the binding preference for polysialic acid</article-title><source>J. Biol. Chem.</source><volume>288</volume><year>2013</year><fpage>33784</fpage><lpage>33796</lpage><pub-id pub-id-type="pmid">24100042</pub-id></element-citation></ref><ref id="bib41"><element-citation publication-type="journal" id="sref41"><person-group person-group-type="author"><name><surname>Naldini</surname><given-names>L.</given-names></name><name><surname>Blömer</surname><given-names>U.</given-names></name><name><surname>Gage</surname><given-names>F.H.</given-names></name><name><surname>Trono</surname><given-names>D.</given-names></name><name><surname>Verma</surname><given-names>I.M.</given-names></name></person-group><article-title>Efficient transfer, integration, and sustained long-term expression of the transgene in adult rat brains injected with a lentiviral vector</article-title><source>Proc. Natl. Acad. Sci. USA</source><volume>93</volume><year>1996</year><fpage>11382</fpage><lpage>11388</lpage><pub-id pub-id-type="pmid">8876144</pub-id></element-citation></ref><ref id="bib42"><element-citation publication-type="journal" id="sref42"><person-group person-group-type="author"><name><surname>Oboho</surname><given-names>I.K.</given-names></name><name><surname>Tomczyk</surname><given-names>S.M.</given-names></name><name><surname>Al-Asmari</surname><given-names>A.M.</given-names></name><name><surname>Banjar</surname><given-names>A.A.</given-names></name><name><surname>Al-Mugti</surname><given-names>H.</given-names></name><name><surname>Aloraini</surname><given-names>M.S.</given-names></name><name><surname>Alkhaldi</surname><given-names>K.Z.</given-names></name><name><surname>Almohammadi</surname><given-names>E.L.</given-names></name><name><surname>Alraddadi</surname><given-names>B.M.</given-names></name><name><surname>Gerber</surname><given-names>S.I.</given-names></name></person-group><article-title>2014 MERS-CoV outbreak in Jeddah--a link to health care facilities</article-title><source>N. Engl. J. Med.</source><volume>372</volume><year>2015</year><fpage>846</fpage><lpage>854</lpage><pub-id pub-id-type="pmid">25714162</pub-id></element-citation></ref><ref id="bib43"><element-citation publication-type="journal" id="sref43"><person-group person-group-type="author"><name><surname>Ogura</surname><given-names>T.</given-names></name><name><surname>Iwasaki</surname><given-names>K.</given-names></name><name><surname>Sato</surname><given-names>C.</given-names></name></person-group><article-title>Topology representing network enables highly accurate classification of protein images taken by cryo electron-microscope without masking</article-title><source>J. Struct. Biol.</source><volume>143</volume><year>2003</year><fpage>185</fpage><lpage>200</lpage><pub-id pub-id-type="pmid">14572474</pub-id></element-citation></ref><ref id="bib44"><element-citation publication-type="journal" id="sref44"><person-group person-group-type="author"><name><surname>Pallesen</surname><given-names>J.</given-names></name><name><surname>Wang</surname><given-names>N.</given-names></name><name><surname>Corbett</surname><given-names>K.S.</given-names></name><name><surname>Wrapp</surname><given-names>D.</given-names></name><name><surname>Kirchdoerfer</surname><given-names>R.N.</given-names></name><name><surname>Turner</surname><given-names>H.L.</given-names></name><name><surname>Cottrell</surname><given-names>C.A.</given-names></name><name><surname>Becker</surname><given-names>M.M.</given-names></name><name><surname>Wang</surname><given-names>L.</given-names></name><name><surname>Shi</surname><given-names>W.</given-names></name></person-group><article-title>Immunogenicity and structures of a rationally designed prefusion MERS-CoV spike antigen</article-title><source>Proc. Natl. Acad. Sci. USA</source><volume>114</volume><year>2017</year><fpage>E7348</fpage><lpage>E7357</lpage><pub-id pub-id-type="pmid">28807998</pub-id></element-citation></ref><ref id="bib45"><element-citation publication-type="journal" id="sref45"><person-group person-group-type="author"><name><surname>Peng</surname><given-names>G.</given-names></name><name><surname>Sun</surname><given-names>D.</given-names></name><name><surname>Rajashankar</surname><given-names>K.R.</given-names></name><name><surname>Qian</surname><given-names>Z.</given-names></name><name><surname>Holmes</surname><given-names>K.V.</given-names></name><name><surname>Li</surname><given-names>F.</given-names></name></person-group><article-title>Crystal structure of mouse coronavirus receptor-binding domain complexed with its murine receptor</article-title><source>Proc. Natl. Acad. Sci. USA</source><volume>108</volume><year>2011</year><fpage>10696</fpage><lpage>10701</lpage><pub-id pub-id-type="pmid">21670291</pub-id></element-citation></ref><ref id="bib46"><element-citation publication-type="journal" id="sref46"><person-group person-group-type="author"><name><surname>Peng</surname><given-names>G.</given-names></name><name><surname>Xu</surname><given-names>L.</given-names></name><name><surname>Lin</surname><given-names>Y.L.</given-names></name><name><surname>Chen</surname><given-names>L.</given-names></name><name><surname>Pasquarella</surname><given-names>J.R.</given-names></name><name><surname>Holmes</surname><given-names>K.V.</given-names></name><name><surname>Li</surname><given-names>F.</given-names></name></person-group><article-title>Crystal structure of bovine coronavirus spike protein lectin domain</article-title><source>J. Biol. Chem.</source><volume>287</volume><year>2012</year><fpage>41931</fpage><lpage>41938</lpage><pub-id pub-id-type="pmid">23091051</pub-id></element-citation></ref><ref id="bib47"><element-citation publication-type="journal" id="sref47"><person-group person-group-type="author"><name><surname>Pettersen</surname><given-names>E.F.</given-names></name><name><surname>Goddard</surname><given-names>T.D.</given-names></name><name><surname>Huang</surname><given-names>C.C.</given-names></name><name><surname>Couch</surname><given-names>G.S.</given-names></name><name><surname>Greenblatt</surname><given-names>D.M.</given-names></name><name><surname>Meng</surname><given-names>E.C.</given-names></name><name><surname>Ferrin</surname><given-names>T.E.</given-names></name></person-group><article-title>UCSF Chimera--a visualization system for exploratory research and analysis</article-title><source>J. Comput. Chem.</source><volume>25</volume><year>2004</year><fpage>1605</fpage><lpage>1612</lpage><pub-id pub-id-type="pmid">15264254</pub-id></element-citation></ref><ref id="bib48"><element-citation publication-type="journal" id="sref48"><person-group person-group-type="author"><name><surname>Potter</surname><given-names>C.S.</given-names></name><name><surname>Chu</surname><given-names>H.</given-names></name><name><surname>Frey</surname><given-names>B.</given-names></name><name><surname>Green</surname><given-names>C.</given-names></name><name><surname>Kisseberth</surname><given-names>N.</given-names></name><name><surname>Madden</surname><given-names>T.J.</given-names></name><name><surname>Miller</surname><given-names>K.L.</given-names></name><name><surname>Nahrstedt</surname><given-names>K.</given-names></name><name><surname>Pulokas</surname><given-names>J.</given-names></name><name><surname>Reilein</surname><given-names>A.</given-names></name></person-group><article-title>Leginon: a system for fully automated acquisition of 1000 electron micrographs a day</article-title><source>Ultramicroscopy</source><volume>77</volume><year>1999</year><fpage>153</fpage><lpage>161</lpage><pub-id pub-id-type="pmid">10406132</pub-id></element-citation></ref><ref id="bib49"><element-citation publication-type="journal" id="sref49"><person-group person-group-type="author"><name><surname>Potterton</surname><given-names>E.</given-names></name><name><surname>Briggs</surname><given-names>P.</given-names></name><name><surname>Turkenburg</surname><given-names>M.</given-names></name><name><surname>Dodson</surname><given-names>E.</given-names></name></person-group><article-title>A graphical user interface to the CCP4 program suite</article-title><source>Acta. Crystallogr. D Biol. Crystallogr.</source><volume>59</volume><year>2003</year><fpage>1131</fpage><lpage>1137</lpage><pub-id pub-id-type="pmid">12832755</pub-id></element-citation></ref><ref id="bib50"><element-citation publication-type="journal" id="sref50"><person-group person-group-type="author"><name><surname>Punjani</surname><given-names>A.</given-names></name><name><surname>Rubinstein</surname><given-names>J.L.</given-names></name><name><surname>Fleet</surname><given-names>D.J.</given-names></name><name><surname>Brubaker</surname><given-names>M.A.</given-names></name></person-group><article-title>cryoSPARC: algorithms for rapid unsupervised cryo-EM structure determination</article-title><source>Nat. Methods</source><volume>14</volume><year>2017</year><fpage>290</fpage><lpage>296</lpage><pub-id pub-id-type="pmid">28165473</pub-id></element-citation></ref><ref id="bib51"><element-citation publication-type="journal" id="sref51"><person-group person-group-type="author"><name><surname>Raj</surname><given-names>V.S.</given-names></name><name><surname>Mou</surname><given-names>H.</given-names></name><name><surname>Smits</surname><given-names>S.L.</given-names></name><name><surname>Dekkers</surname><given-names>D.H.</given-names></name><name><surname>Müller</surname><given-names>M.A.</given-names></name><name><surname>Dijkman</surname><given-names>R.</given-names></name><name><surname>Muth</surname><given-names>D.</given-names></name><name><surname>Demmers</surname><given-names>J.A.</given-names></name><name><surname>Zaki</surname><given-names>A.</given-names></name><name><surname>Fouchier</surname><given-names>R.A.</given-names></name></person-group><article-title>Dipeptidyl peptidase 4 is a functional receptor for the emerging human coronavirus-EMC</article-title><source>Nature</source><volume>495</volume><year>2013</year><fpage>251</fpage><lpage>254</lpage><pub-id pub-id-type="pmid">23486063</pub-id></element-citation></ref><ref id="bib52"><element-citation publication-type="journal" id="sref52"><person-group person-group-type="author"><name><surname>Rockx</surname><given-names>B.</given-names></name><name><surname>Corti</surname><given-names>D.</given-names></name><name><surname>Donaldson</surname><given-names>E.</given-names></name><name><surname>Sheahan</surname><given-names>T.</given-names></name><name><surname>Stadler</surname><given-names>K.</given-names></name><name><surname>Lanzavecchia</surname><given-names>A.</given-names></name><name><surname>Baric</surname><given-names>R.</given-names></name></person-group><article-title>Structural basis for potent cross-neutralizing human monoclonal antibody protection against lethal human and zoonotic severe acute respiratory syndrome coronavirus challenge</article-title><source>J. Virol.</source><volume>82</volume><year>2008</year><fpage>3220</fpage><lpage>3235</lpage><pub-id pub-id-type="pmid">18199635</pub-id></element-citation></ref><ref id="bib53"><element-citation publication-type="journal" id="sref53"><person-group person-group-type="author"><name><surname>Rosen</surname><given-names>O.</given-names></name><name><surname>Chan</surname><given-names>L.L.</given-names></name><name><surname>Abiona</surname><given-names>O.M.</given-names></name><name><surname>Gough</surname><given-names>P.</given-names></name><name><surname>Wang</surname><given-names>L.</given-names></name><name><surname>Shi</surname><given-names>W.</given-names></name><name><surname>Zhang</surname><given-names>Y.</given-names></name><name><surname>Wang</surname><given-names>N.</given-names></name><name><surname>Kong</surname><given-names>W.P.</given-names></name><name><surname>McLellan</surname><given-names>J.S.</given-names></name></person-group><article-title>A high-throughput inhibition assay to study MERS-CoV antibody interactions using image cytometry</article-title><source>J. Virol. Methods</source><volume>265</volume><year>2019</year><fpage>77</fpage><lpage>83</lpage><pub-id pub-id-type="pmid">30468747</pub-id></element-citation></ref><ref id="bib54"><element-citation publication-type="journal" id="sref54"><person-group person-group-type="author"><name><surname>Schultze</surname><given-names>B.</given-names></name><name><surname>Gross</surname><given-names>H.J.</given-names></name><name><surname>Brossmer</surname><given-names>R.</given-names></name><name><surname>Herrler</surname><given-names>G.</given-names></name></person-group><article-title>The S protein of bovine coronavirus is a hemagglutinin recognizing 9-O-acetylated sialic acid as a receptor determinant</article-title><source>J. Virol.</source><volume>65</volume><year>1991</year><fpage>6232</fpage><lpage>6237</lpage><pub-id pub-id-type="pmid">1920630</pub-id></element-citation></ref><ref id="bib55"><element-citation publication-type="journal" id="sref55"><person-group person-group-type="author"><name><surname>Scobey</surname><given-names>T.</given-names></name><name><surname>Yount</surname><given-names>B.L.</given-names></name><name><surname>Sims</surname><given-names>A.C.</given-names></name><name><surname>Donaldson</surname><given-names>E.F.</given-names></name><name><surname>Agnihothram</surname><given-names>S.S.</given-names></name><name><surname>Menachery</surname><given-names>V.D.</given-names></name><name><surname>Graham</surname><given-names>R.L.</given-names></name><name><surname>Swanstrom</surname><given-names>J.</given-names></name><name><surname>Bove</surname><given-names>P.F.</given-names></name><name><surname>Kim</surname><given-names>J.D.</given-names></name></person-group><article-title>Reverse genetics with a full-length infectious cDNA of the Middle East respiratory syndrome coronavirus</article-title><source>Proc. Natl. Acad. Sci. USA</source><volume>110</volume><year>2013</year><fpage>16157</fpage><lpage>16162</lpage><pub-id pub-id-type="pmid">24043791</pub-id></element-citation></ref><ref id="bib56"><element-citation publication-type="journal" id="sref56"><person-group person-group-type="author"><name><surname>Shang</surname><given-names>J.</given-names></name><name><surname>Zheng</surname><given-names>Y.</given-names></name><name><surname>Yang</surname><given-names>Y.</given-names></name><name><surname>Liu</surname><given-names>C.</given-names></name><name><surname>Geng</surname><given-names>Q.</given-names></name><name><surname>Luo</surname><given-names>C.</given-names></name><name><surname>Zhang</surname><given-names>W.</given-names></name><name><surname>Li</surname><given-names>F.</given-names></name></person-group><article-title>Cryo-EM structure of infectious bronchitis coronavirus spike protein reveals structural and functional evolution of coronavirus spike proteins</article-title><source>PLoS Pathog.</source><volume>14</volume><year>2018</year><fpage>e1007009</fpage><pub-id pub-id-type="pmid">29684066</pub-id></element-citation></ref><ref id="bib57"><element-citation publication-type="journal" id="sref57"><person-group person-group-type="author"><name><surname>Shang</surname><given-names>J.</given-names></name><name><surname>Zheng</surname><given-names>Y.</given-names></name><name><surname>Yang</surname><given-names>Y.</given-names></name><name><surname>Liu</surname><given-names>C.</given-names></name><name><surname>Geng</surname><given-names>Q.</given-names></name><name><surname>Tai</surname><given-names>W.</given-names></name><name><surname>Du</surname><given-names>L.</given-names></name><name><surname>Zhou</surname><given-names>Y.</given-names></name><name><surname>Zhang</surname><given-names>W.</given-names></name><name><surname>Li</surname><given-names>F.</given-names></name></person-group><article-title>Cryo-Electron Microscopy Structure of Porcine Deltacoronavirus Spike Protein in the Prefusion State</article-title><source>J. Virol.</source><volume>92</volume><year>2018</year><fpage>e01556-17</fpage><pub-id pub-id-type="pmid">29070693</pub-id></element-citation></ref><ref id="bib58"><element-citation publication-type="journal" id="sref58"><person-group person-group-type="author"><name><surname>Song</surname><given-names>W.</given-names></name><name><surname>Gui</surname><given-names>M.</given-names></name><name><surname>Wang</surname><given-names>X.</given-names></name><name><surname>Xiang</surname><given-names>Y.</given-names></name></person-group><article-title>Cryo-EM structure of the SARS coronavirus spike glycoprotein in complex with its host cell receptor ACE2</article-title><source>PLoS Pathog.</source><volume>14</volume><year>2018</year><fpage>e1007236</fpage><pub-id pub-id-type="pmid">30102747</pub-id></element-citation></ref><ref id="bib59"><element-citation publication-type="journal" id="sref59"><person-group person-group-type="author"><name><surname>Stewart</surname><given-names>S.A.</given-names></name><name><surname>Dykxhoorn</surname><given-names>D.M.</given-names></name><name><surname>Palliser</surname><given-names>D.</given-names></name><name><surname>Mizuno</surname><given-names>H.</given-names></name><name><surname>Yu</surname><given-names>E.Y.</given-names></name><name><surname>An</surname><given-names>D.S.</given-names></name><name><surname>Sabatini</surname><given-names>D.M.</given-names></name><name><surname>Chen</surname><given-names>I.S.</given-names></name><name><surname>Hahn</surname><given-names>W.C.</given-names></name><name><surname>Sharp</surname><given-names>P.A.</given-names></name></person-group><article-title>Lentivirus-delivered stable gene silencing by RNAi in primary cells</article-title><source>RNA</source><volume>9</volume><year>2003</year><fpage>493</fpage><lpage>501</lpage><pub-id pub-id-type="pmid">12649500</pub-id></element-citation></ref><ref id="bib60"><element-citation publication-type="journal" id="sref60"><person-group person-group-type="author"><name><surname>Suloway</surname><given-names>C.</given-names></name><name><surname>Pulokas</surname><given-names>J.</given-names></name><name><surname>Fellmann</surname><given-names>D.</given-names></name><name><surname>Cheng</surname><given-names>A.</given-names></name><name><surname>Guerra</surname><given-names>F.</given-names></name><name><surname>Quispe</surname><given-names>J.</given-names></name><name><surname>Stagg</surname><given-names>S.</given-names></name><name><surname>Potter</surname><given-names>C.S.</given-names></name><name><surname>Carragher</surname><given-names>B.</given-names></name></person-group><article-title>Automated molecular microscopy: the new Leginon system</article-title><source>J. Struct. Biol.</source><volume>151</volume><year>2005</year><fpage>41</fpage><lpage>60</lpage><pub-id pub-id-type="pmid">15890530</pub-id></element-citation></ref><ref id="bib61"><element-citation publication-type="journal" id="sref61"><person-group person-group-type="author"><name><surname>Tang</surname><given-names>G.</given-names></name><name><surname>Peng</surname><given-names>L.</given-names></name><name><surname>Baldwin</surname><given-names>P.R.</given-names></name><name><surname>Mann</surname><given-names>D.S.</given-names></name><name><surname>Jiang</surname><given-names>W.</given-names></name><name><surname>Rees</surname><given-names>I.</given-names></name><name><surname>Ludtke</surname><given-names>S.J.</given-names></name></person-group><article-title>EMAN2: an extensible image processing suite for electron microscopy</article-title><source>J. Struct. Biol.</source><volume>157</volume><year>2007</year><fpage>38</fpage><lpage>46</lpage><pub-id pub-id-type="pmid">16859925</pub-id></element-citation></ref><ref id="bib62"><element-citation publication-type="journal" id="sref62"><person-group person-group-type="author"><name><surname>Tang</surname><given-names>X.C.</given-names></name><name><surname>Agnihothram</surname><given-names>S.S.</given-names></name><name><surname>Jiao</surname><given-names>Y.</given-names></name><name><surname>Stanhope</surname><given-names>J.</given-names></name><name><surname>Graham</surname><given-names>R.L.</given-names></name><name><surname>Peterson</surname><given-names>E.C.</given-names></name><name><surname>Avnir</surname><given-names>Y.</given-names></name><name><surname>Tallarico</surname><given-names>A.S.</given-names></name><name><surname>Sheehan</surname><given-names>J.</given-names></name><name><surname>Zhu</surname><given-names>Q.</given-names></name></person-group><article-title>Identification of human neutralizing antibodies against MERS-CoV and their role in virus adaptive evolution</article-title><source>Proc. Natl. Acad. Sci. USA</source><volume>111</volume><year>2014</year><fpage>E2018</fpage><lpage>E2026</lpage><pub-id pub-id-type="pmid">24778221</pub-id></element-citation></ref><ref id="bib63"><element-citation publication-type="journal" id="sref63"><person-group person-group-type="author"><name><surname>van Boheemen</surname><given-names>S.</given-names></name><name><surname>de Graaf</surname><given-names>M.</given-names></name><name><surname>Lauber</surname><given-names>C.</given-names></name><name><surname>Bestebroer</surname><given-names>T.M.</given-names></name><name><surname>Raj</surname><given-names>V.S.</given-names></name><name><surname>Zaki</surname><given-names>A.M.</given-names></name><name><surname>Osterhaus</surname><given-names>A.D.</given-names></name><name><surname>Haagmans</surname><given-names>B.L.</given-names></name><name><surname>Gorbalenya</surname><given-names>A.E.</given-names></name><name><surname>Snijder</surname><given-names>E.J.</given-names></name><name><surname>Fouchier</surname><given-names>R.A.</given-names></name></person-group><article-title>Genomic characterization of a newly discovered coronavirus associated with acute respiratory distress syndrome in humans</article-title><source>MBio</source><volume>3</volume><year>2012</year><comment>e00473-12</comment></element-citation></ref><ref id="bib64"><element-citation publication-type="journal" id="sref64"><person-group person-group-type="author"><name><surname>Voss</surname><given-names>N.R.</given-names></name><name><surname>Yoshioka</surname><given-names>C.K.</given-names></name><name><surname>Radermacher</surname><given-names>M.</given-names></name><name><surname>Potter</surname><given-names>C.S.</given-names></name><name><surname>Carragher</surname><given-names>B.</given-names></name></person-group><article-title>DoG Picker and TiltPicker: software tools to facilitate particle selection in single particle electron microscopy</article-title><source>J. Struct. Biol.</source><volume>166</volume><year>2009</year><fpage>205</fpage><lpage>213</lpage><pub-id pub-id-type="pmid">19374019</pub-id></element-citation></ref><ref id="bib65"><element-citation publication-type="journal" id="sref65"><person-group person-group-type="author"><name><surname>Walls</surname><given-names>A.C.</given-names></name><name><surname>Tortorici</surname><given-names>M.A.</given-names></name><name><surname>Bosch</surname><given-names>B.J.</given-names></name><name><surname>Frenz</surname><given-names>B.</given-names></name><name><surname>Rottier</surname><given-names>P.J.M.</given-names></name><name><surname>DiMaio</surname><given-names>F.</given-names></name><name><surname>Rey</surname><given-names>F.A.</given-names></name><name><surname>Veesler</surname><given-names>D.</given-names></name></person-group><article-title>Cryo-electron microscopy structure of a coronavirus spike glycoprotein trimer</article-title><source>Nature</source><volume>531</volume><year>2016</year><fpage>114</fpage><lpage>117</lpage><pub-id pub-id-type="pmid">26855426</pub-id></element-citation></ref><ref id="bib66"><element-citation publication-type="journal" id="sref66"><person-group person-group-type="author"><name><surname>Walls</surname><given-names>A.C.</given-names></name><name><surname>Tortorici</surname><given-names>M.A.</given-names></name><name><surname>Frenz</surname><given-names>B.</given-names></name><name><surname>Snijder</surname><given-names>J.</given-names></name><name><surname>Li</surname><given-names>W.</given-names></name><name><surname>Rey</surname><given-names>F.A.</given-names></name><name><surname>DiMaio</surname><given-names>F.</given-names></name><name><surname>Bosch</surname><given-names>B.J.</given-names></name><name><surname>Veesler</surname><given-names>D.</given-names></name></person-group><article-title>Glycan shield and epitope masking of a coronavirus spike protein observed by cryo-electron microscopy</article-title><source>Nat. Struct. Mol. Biol.</source><volume>23</volume><year>2016</year><fpage>899</fpage><lpage>905</lpage><pub-id pub-id-type="pmid">27617430</pub-id></element-citation></ref><ref id="bib67"><element-citation publication-type="journal" id="sref67"><person-group person-group-type="author"><name><surname>Walls</surname><given-names>A.C.</given-names></name><name><surname>Xiong</surname><given-names>X.</given-names></name><name><surname>Park</surname><given-names>Y.J.</given-names></name><name><surname>Tortorici</surname><given-names>M.A.</given-names></name><name><surname>Snijder</surname><given-names>J.</given-names></name><name><surname>Quispe</surname><given-names>J.</given-names></name><name><surname>Cameroni</surname><given-names>E.</given-names></name><name><surname>Gopal</surname><given-names>R.</given-names></name><name><surname>Dai</surname><given-names>M.</given-names></name><name><surname>Lanzavecchia</surname><given-names>A.</given-names></name></person-group><article-title>Unexpected Receptor Functional Mimicry Elucidates Activation of Coronavirus Fusion</article-title><source>Cell</source><volume>176</volume><year>2019</year><fpage>1026</fpage><lpage>1039.e15</lpage><pub-id pub-id-type="pmid">30712865</pub-id></element-citation></ref><ref id="bib68"><element-citation publication-type="journal" id="sref68"><person-group person-group-type="author"><name><surname>Wang</surname><given-names>L.</given-names></name><name><surname>Shi</surname><given-names>W.</given-names></name><name><surname>Joyce</surname><given-names>M.G.</given-names></name><name><surname>Modjarrad</surname><given-names>K.</given-names></name><name><surname>Zhang</surname><given-names>Y.</given-names></name><name><surname>Leung</surname><given-names>K.</given-names></name><name><surname>Lees</surname><given-names>C.R.</given-names></name><name><surname>Zhou</surname><given-names>T.</given-names></name><name><surname>Yassine</surname><given-names>H.M.</given-names></name><name><surname>Kanekiyo</surname><given-names>M.</given-names></name></person-group><article-title>Evaluation of candidate vaccine approaches for MERS-CoV</article-title><source>Nat. Commun.</source><volume>6</volume><year>2015</year><fpage>7712</fpage><pub-id pub-id-type="pmid">26218507</pub-id></element-citation></ref><ref id="bib69"><element-citation publication-type="journal" id="sref69"><person-group person-group-type="author"><name><surname>Wang</surname><given-names>L.</given-names></name><name><surname>Shi</surname><given-names>W.</given-names></name><name><surname>Chappell</surname><given-names>J.D.</given-names></name><name><surname>Joyce</surname><given-names>M.G.</given-names></name><name><surname>Zhang</surname><given-names>Y.</given-names></name><name><surname>Kanekiyo</surname><given-names>M.</given-names></name><name><surname>Becker</surname><given-names>M.M.</given-names></name><name><surname>van Doremalen</surname><given-names>N.</given-names></name><name><surname>Fischer</surname><given-names>R.</given-names></name><name><surname>Wang</surname><given-names>N.</given-names></name></person-group><article-title>Importance of Neutralizing Monoclonal Antibodies Targeting Multiple Antigenic Sites on the Middle East Respiratory Syndrome Coronavirus Spike Glycoprotein To Avoid Neutralization Escape</article-title><source>J. Virol.</source><volume>92</volume><year>2018</year><fpage>e02002-17</fpage><pub-id pub-id-type="pmid">29514901</pub-id></element-citation></ref><ref id="bib70"><element-citation publication-type="journal" id="sref70"><person-group person-group-type="author"><name><surname>Wang</surname><given-names>N.</given-names></name><name><surname>Shi</surname><given-names>X.</given-names></name><name><surname>Jiang</surname><given-names>L.</given-names></name><name><surname>Zhang</surname><given-names>S.</given-names></name><name><surname>Wang</surname><given-names>D.</given-names></name><name><surname>Tong</surname><given-names>P.</given-names></name><name><surname>Guo</surname><given-names>D.</given-names></name><name><surname>Fu</surname><given-names>L.</given-names></name><name><surname>Cui</surname><given-names>Y.</given-names></name><name><surname>Liu</surname><given-names>X.</given-names></name></person-group><article-title>Structure of MERS-CoV spike receptor-binding domain complexed with human receptor DPP4</article-title><source>Cell Res.</source><volume>23</volume><year>2013</year><fpage>986</fpage><lpage>993</lpage><pub-id pub-id-type="pmid">23835475</pub-id></element-citation></ref><ref id="bib71"><element-citation publication-type="book" id="sref71"><person-group person-group-type="author"><collab>WHO</collab></person-group><chapter-title>Middle East respiratory syndrome coronavirus (MERS-CoV)</chapter-title><year>2018</year><publisher-name>World Health Organization</publisher-name><ext-link ext-link-type="uri" xlink:href="https://www.who.int/emergencies/mers-cov/en/" id="intref0175">https://www.who.int/emergencies/mers-cov/en/</ext-link></element-citation></ref><ref id="bib72"><element-citation publication-type="journal" id="sref72"><person-group person-group-type="author"><name><surname>Xiong</surname><given-names>X.</given-names></name><name><surname>Tortorici</surname><given-names>M.A.</given-names></name><name><surname>Snijder</surname><given-names>J.</given-names></name><name><surname>Yoshioka</surname><given-names>C.</given-names></name><name><surname>Walls</surname><given-names>A.C.</given-names></name><name><surname>Li</surname><given-names>W.</given-names></name><name><surname>McGuire</surname><given-names>A.T.</given-names></name><name><surname>Rey</surname><given-names>F.A.</given-names></name><name><surname>Bosch</surname><given-names>B.J.</given-names></name><name><surname>Veesler</surname><given-names>D.</given-names></name></person-group><article-title>Glycan Shield and Fusion Activation of a Deltacoronavirus Spike Glycoprotein Fine-Tuned for Enteric Infections</article-title><source>J. Virol.</source><volume>92</volume><year>2018</year><fpage>e01628-17</fpage><pub-id pub-id-type="pmid">29093093</pub-id></element-citation></ref><ref id="bib73"><element-citation publication-type="journal" id="sref73"><person-group person-group-type="author"><name><surname>Ying</surname><given-names>T.</given-names></name><name><surname>Du</surname><given-names>L.</given-names></name><name><surname>Ju</surname><given-names>T.W.</given-names></name><name><surname>Prabakaran</surname><given-names>P.</given-names></name><name><surname>Lau</surname><given-names>C.C.</given-names></name><name><surname>Lu</surname><given-names>L.</given-names></name><name><surname>Liu</surname><given-names>Q.</given-names></name><name><surname>Wang</surname><given-names>L.</given-names></name><name><surname>Feng</surname><given-names>Y.</given-names></name><name><surname>Wang</surname><given-names>Y.</given-names></name></person-group><article-title>Exceptionally potent neutralization of Middle East respiratory syndrome coronavirus by human monoclonal antibodies</article-title><source>J. Virol.</source><volume>88</volume><year>2014</year><fpage>7796</fpage><lpage>7805</lpage><pub-id pub-id-type="pmid">24789777</pub-id></element-citation></ref><ref id="bib74"><element-citation publication-type="journal" id="sref74"><person-group person-group-type="author"><name><surname>Yuan</surname><given-names>Y.</given-names></name><name><surname>Cao</surname><given-names>D.</given-names></name><name><surname>Zhang</surname><given-names>Y.</given-names></name><name><surname>Ma</surname><given-names>J.</given-names></name><name><surname>Qi</surname><given-names>J.</given-names></name><name><surname>Wang</surname><given-names>Q.</given-names></name><name><surname>Lu</surname><given-names>G.</given-names></name><name><surname>Wu</surname><given-names>Y.</given-names></name><name><surname>Yan</surname><given-names>J.</given-names></name><name><surname>Shi</surname><given-names>Y.</given-names></name></person-group><article-title>Cryo-EM structures of MERS-CoV and SARS-CoV spike glycoproteins reveal the dynamic receptor binding domains</article-title><source>Nat. Commun.</source><volume>8</volume><year>2017</year><fpage>15092</fpage><pub-id pub-id-type="pmid">28393837</pub-id></element-citation></ref><ref id="bib75"><element-citation publication-type="journal" id="sref75"><person-group person-group-type="author"><name><surname>Yusof</surname><given-names>M.F.</given-names></name><name><surname>Queen</surname><given-names>K.</given-names></name><name><surname>Eltahir</surname><given-names>Y.M.</given-names></name><name><surname>Paden</surname><given-names>C.R.</given-names></name><name><surname>Al Hammadi</surname><given-names>Z.M.A.H.</given-names></name><name><surname>Tao</surname><given-names>Y.</given-names></name><name><surname>Li</surname><given-names>Y.</given-names></name><name><surname>Khalafalla</surname><given-names>A.I.</given-names></name><name><surname>Shi</surname><given-names>M.</given-names></name><name><surname>Zhang</surname><given-names>J.</given-names></name></person-group><article-title>Diversity of Middle East respiratory syndrome coronaviruses in 109 dromedary camels based on full-genome sequencing, Abu Dhabi, United Arab Emirates</article-title><source>Emerg. Microbes Infect.</source><volume>6</volume><year>2017</year><fpage>e101</fpage><pub-id pub-id-type="pmid">29116217</pub-id></element-citation></ref><ref id="bib76"><element-citation publication-type="journal" id="sref76"><person-group person-group-type="author"><name><surname>Zaki</surname><given-names>A.M.</given-names></name><name><surname>van Boheemen</surname><given-names>S.</given-names></name><name><surname>Bestebroer</surname><given-names>T.M.</given-names></name><name><surname>Osterhaus</surname><given-names>A.D.</given-names></name><name><surname>Fouchier</surname><given-names>R.A.</given-names></name></person-group><article-title>Isolation of a novel coronavirus from a man with pneumonia in Saudi Arabia</article-title><source>N. Engl. J. Med.</source><volume>367</volume><year>2012</year><fpage>1814</fpage><lpage>1820</lpage><pub-id pub-id-type="pmid">23075143</pub-id></element-citation></ref><ref id="bib77"><element-citation publication-type="journal" id="sref77"><person-group person-group-type="author"><name><surname>Zhang</surname><given-names>K.</given-names></name></person-group><article-title>Gctf: Real-time CTF determination and correction</article-title><source>J. Struct. Biol.</source><volume>193</volume><year>2016</year><fpage>1</fpage><lpage>12</lpage><pub-id pub-id-type="pmid">26592709</pub-id></element-citation></ref><ref id="bib78"><element-citation publication-type="journal" id="sref78"><person-group person-group-type="author"><name><surname>Zheng</surname><given-names>S.Q.</given-names></name><name><surname>Palovcak</surname><given-names>E.</given-names></name><name><surname>Armache</surname><given-names>J.P.</given-names></name><name><surname>Verba</surname><given-names>K.A.</given-names></name><name><surname>Cheng</surname><given-names>Y.</given-names></name><name><surname>Agard</surname><given-names>D.A.</given-names></name></person-group><article-title>MotionCor2: anisotropic correction of beam-induced motion for improved cryo-electron microscopy</article-title><source>Nat. Methods</source><volume>14</volume><year>2017</year><fpage>331</fpage><lpage>332</lpage><pub-id pub-id-type="pmid">28250466</pub-id></element-citation></ref><ref id="bib79"><element-citation publication-type="journal" id="sref79"><person-group person-group-type="author"><name><surname>Zivanov</surname><given-names>J.</given-names></name><name><surname>Nakane</surname><given-names>T.</given-names></name><name><surname>Forsberg</surname><given-names>B.O.</given-names></name><name><surname>Kimanius</surname><given-names>D.</given-names></name><name><surname>Hagen</surname><given-names>W.J.</given-names></name><name><surname>Lindahl</surname><given-names>E.</given-names></name><name><surname>Scheres</surname><given-names>S.H.</given-names></name></person-group><article-title>New tools for automated high-resolution cryo-EM structure determination in RELION-3</article-title><source>eLife</source><volume>7</volume><year>2018</year><fpage>e42166</fpage><pub-id pub-id-type="pmid">30412051</pub-id></element-citation></ref></ref-list><sec id="app2" sec-type="supplementary-material"><title>Supplemental Information</title><p id="p0375"><supplementary-material content-type="local-data" id="mmc1"><caption><title>Document S1. Figures S1–S3 and Tables S1–S3</title></caption><media xlink:href="mmc1.pdf"></media></supplementary-material><supplementary-material content-type="local-data" id="mmc2"><caption><title>Document S2. Article plus Supplemental Information</title></caption><media xlink:href="mmc2.pdf"></media></supplementary-material></p></sec><ack id="ack0010"><title>Acknowledgments</title><p>We thank colleagues and members of our labs for comments on the manuscript, Emilie Shipman (Dartmouth College) and John Ludes-Meyers (UT-Austin) for assistance with protein expression, the beamline scientists for X-ray data collection support at SBC 19-ID (Argonne National Laboratory), Xiaotao Lu (Vanderbilt) for mutant virus construction using the MERS-CoV BAC, Erica Andres (Vanderbilt) and Xiaotao Lu for assistance with selection and sequence analysis of G2 escape-mutant viruses, and Gabriel Ozorowski for assistance with cryo-EM model building and refinement. This work was supported by NIH grant R01AI127521 (to J.S.M. and A.B.W.), <funding-source id="gs1">NIH</funding-source> contract HHSN261200800001E agreement 16x142 (to M.R.D. and J.D.C.), and intramural funding from <funding-source id="gs2">National Institute of Allergy and Infectious Diseases</funding-source> for work at the VRC (B.S.G.). Argonne is operated by UChicago Argonne, LLC, for the <funding-source id="gs3">U.S. Department of Energy</funding-source> (DOE), <funding-source id="gs4">Office of Biological and Environmental Research</funding-source> under contract DE-AC02-06CH11357.</p><sec id="sec5"><title>Author Contributions</title><p id="p0360">N.W. designed and produced MERS-CoV S, S1-NTD, and G2 Fab proteins and different variants; crystallized G2 Fab and NTD–Fab complex and determined the structures; and conducted the SPR assay, cell-surface binding assay, and competition assay. O.R. and K.S.C. conducted the study to compare the efficacy of G2 Fab and IgG on binding inhibition and neutralization. L.W. conducted the neutralizing assay of G2 against pseudoviruses bearing MERS-CoV S, MERS-CoV S-S28F, and MERS-CoV S-G198D. W.S., Y.Z., and K.L. generated reagents used in multiple assays. H.L.T. performed the negative-stain EM study. J.P. collected the cryo-EM data and C.A.B. processed the data and determined the structure. M.M.B., J.D.C., and M.R.D. isolated and analyzed G2-escape variants of MERS-CoV. L.J.S., J.D.C., and M.R.D. conducted neutralization studies with authentic MERS-CoV (or MERS-CoV variants) in a BSL-3 setting. N.W., A.B.W., B.S.G, and J.S.M. conceived and designed the study and analyzed data. N.W. and J.S.M wrote the initial draft of the manuscript, on which all authors provided edits and comments.</p></sec><sec sec-type="COI-statement" id="sec6"><title>Declaration of Interests</title><p id="p0365">N.W., K.S.C., R.N.K., H.L.T., B.S.G., A.B.W., and J.S.M. are inventors on U.S. patent application 62/412,703, entitled “Prefusion Coronavirus Spike Proteins and Their Use.” L.W., W.S., and B.S.G. are inventors on U.S. patent application PCT/US2016/019395, entitled “Middle East Respiratory Syndrome Coronavirus Immunogens, Antibodies and Their Use.”</p></sec></ack><fn-group><fn id="app1" fn-type="supplementary-material"><p id="p0370">Supplemental Information can be found online at <ext-link ext-link-type="uri" xlink:href="https://doi.org/10.1016/j.celrep.2019.08.052" id="intref0170">https://doi.org/10.1016/j.celrep.2019.08.052</ext-link>.</p></fn></fn-group></back></pmc></record>Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Sante/explor/MersV1/Data/Pmc/Corpus
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 000B58 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Pmc/Corpus/biblio.hfd -nk 000B58 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien
|wiki= Sante
|area= MersV1
|flux= Pmc
|étape= Corpus
|type= RBID
|clé=
|texte=
}}
| This area was generated with Dilib version V0.6.33. |  |