Deducing the Crystal Structure of MERS-CoV Helicase.
Identifieur interne : 000744 ( 2020/Analysis ); précédent : 000743; suivant : 000745Deducing the Crystal Structure of MERS-CoV Helicase.
Auteurs : Sheng Cui [République populaire de Chine] ; Wei Hao [République populaire de Chine]Source :
- Methods in molecular biology (Clifton, N.J.) [ 1940-6029 ] ; 2020.
Abstract
RNA virus encodes a helicase essential for viral RNA transcription and replication when the genome size is larger than 7 kb. Coronavirus (CoV) has an exceptionally large RNA genome (~30 kb) and it encodes an essential replicase, the nonstructural protein 13 (nsp13), a member of superfamily 1 helicases. Nsp13 is among the evolutionary most conserved proteins not only in CoVs but also in nidovirales. Thus, it is considered as an important drug target. However, the high-resolution structure of CoV nsp13 remained unavailable even until more than a decade after the outbreak of the severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003, which hindered the structure-based drug design. This is in part due to the intrinsic flexibility of nsp13. Here, we describe protocols of deducing the crystal structure of Middle East respiratory syndrome coronavirus (MERS-CoV) helicase in detail, which include protein expression, purification, crystallization, enzymatic characterization, and structure determination. With these methods, catalytically active recombinant MERS-CoV nsp13 protein can be prepared and crystallized and the crystal structure can be solved.
DOI: 10.1007/978-1-0716-0211-9_6
PubMed: 31883088
Affiliations:
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R. China. Cui.sheng@ipb.pumc.edu.cn.</nlm:affiliation><country wicri:rule="url">République populaire de Chine</country></affiliation></author><author><name sortKey="Hao, Wei" sort="Hao, Wei" uniqKey="Hao W" first="Wei" last="Hao">Wei Hao</name><affiliation wicri:level="3"><nlm:affiliation>NHC Key Laboratory of Systems Biology of Pathogens, Institute of Pathogen Biology, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, P. R. China.</nlm:affiliation><country xml:lang="fr" wicri:curation="lc">République populaire de Chine</country><wicri:regionArea>NHC Key Laboratory of Systems Biology of Pathogens, Institute of Pathogen Biology, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing</wicri:regionArea><placeName><settlement type="city">Pékin</settlement></placeName></affiliation></author></analytic><series><title level="j">Methods in molecular biology (Clifton, N.J.)</title><idno type="eISSN">1940-6029</idno><imprint><date when="2020" type="published">2020</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">RNA virus encodes a helicase essential for viral RNA transcription and replication when the genome size is larger than 7 kb. Coronavirus (CoV) has an exceptionally large RNA genome (~30 kb) and it encodes an essential replicase, the nonstructural protein 13 (nsp13), a member of superfamily 1 helicases. Nsp13 is among the evolutionary most conserved proteins not only in CoVs but also in nidovirales. Thus, it is considered as an important drug target. However, the high-resolution structure of CoV nsp13 remained unavailable even until more than a decade after the outbreak of the severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003, which hindered the structure-based drug design. This is in part due to the intrinsic flexibility of nsp13. Here, we describe protocols of deducing the crystal structure of Middle East respiratory syndrome coronavirus (MERS-CoV) helicase in detail, which include protein expression, purification, crystallization, enzymatic characterization, and structure determination. With these methods, catalytically active recombinant MERS-CoV nsp13 protein can be prepared and crystallized and the crystal structure can be solved.</div></front></TEI><affiliations><list><country><li>République populaire de Chine</li></country><settlement><li>Pékin</li></settlement></list><tree><country name="République populaire de Chine"><noRegion><name sortKey="Cui, Sheng" sort="Cui, Sheng" uniqKey="Cui S" first="Sheng" last="Cui">Sheng Cui</name></noRegion><name sortKey="Hao, Wei" sort="Hao, Wei" uniqKey="Hao W" first="Wei" last="Hao">Wei Hao</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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