Biochemical characterization of a recombinant SARS coronavirus nsp12 RNA-dependent RNA polymerase capable of copying viral RNA templates
Identifieur interne : 000949 ( PascalFrancis/Curation ); précédent : 000948; suivant : 000950Biochemical characterization of a recombinant SARS coronavirus nsp12 RNA-dependent RNA polymerase capable of copying viral RNA templates
Auteurs : Dae-Gyun Ahn [Corée du Sud] ; Jin-Kyu Choi [Corée du Sud] ; Deborah R. Taylor [États-Unis] ; Jong-Won Oh [Corée du Sud]Source :
- Archives of virology [ 0304-8608 ] ; 2012.
Descripteurs français
- Pascal (Inist)
English descriptors
Abstract
The severe acute respiratory syndrome coronavirus (SARS-CoV) RNA genome is replicated by a virus-encoded RNA replicase, the key component of which is the nonstructural protein 12 (nsp12). In this report, we describe the biochemical properties of a full-length recombinant SARS-CoV nsp12 RNA-dependent RNA polymerase (RdRp) capable of copying viral RNA templates. The purified SARS-CoV nsp12 showed both primer-dependent and primer-independent RNA synthesis activities using homopolymeric RNA templates. The RdRp activity was strictly dependent on Mn2+. The nsp12 preferentially copied homopolymeric pyrimidine RNA templates in the absence of an added oligonucleotide primer. It was also able to initiate de novo RNA synthesis from the 3'-ends of both the plus- and minus-strand genome of SARS-CoV, using the 3'-terminal 36- and 37-nt RNA, respectively. The in vitro RdRp assay system established with a full-length nsp12 will be useful for understanding the mechanisms of coronavirus replication and for the development of anti-SARS-CoV agents.
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Taylor</name><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Laboratory of Emerging Pathogens, Division of Emerging and Transfusion Transmitted Diseases, Office of Blood Research and Review, Center for Biologics Evaluation and Research, Food and Drug Administration</s1><s2>Bethesda, MD 20892</s2><s3>USA</s3><sZ>3 aut.</sZ></inist:fA14><country>États-Unis</country></affiliation></author><author><name sortKey="Oh, Jong Won" sort="Oh, Jong Won" uniqKey="Oh J" first="Jong-Won" last="Oh">Jong-Won Oh</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Biotechnology, Yonsei University, 134 Shinchon-dong</s1><s2>Seodaemun-gu, Seoul 120-749</s2><s3>KOR</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ></inist:fA14><country>Corée du Sud</country></affiliation><affiliation wicri:level="1"><inist:fA14 i1="03"><s1>Translational Research Center for Protein Function Control, Yonsei University, 134 Shinchon-dong</s1><s2>Seodaemun-gu, Seoul 120-749</s2><s3>KOR</s3><sZ>4 aut.</sZ></inist:fA14><country>Corée du Sud</country></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">INIST</idno><idno type="inist">12-0437260</idno><date when="2012">2012</date><idno type="stanalyst">PASCAL 12-0437260 INIST</idno><idno type="RBID">Pascal:12-0437260</idno><idno type="wicri:Area/PascalFrancis/Corpus">000039</idno><idno type="wicri:Area/PascalFrancis/Curation">000949</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a">Biochemical characterization of a recombinant SARS coronavirus nsp12 RNA-dependent RNA polymerase capable of copying viral RNA templates</title><author><name sortKey="Ahn, Dae Gyun" sort="Ahn, Dae Gyun" uniqKey="Ahn D" first="Dae-Gyun" last="Ahn">Dae-Gyun Ahn</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Biotechnology, Yonsei University, 134 Shinchon-dong</s1><s2>Seodaemun-gu, Seoul 120-749</s2><s3>KOR</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ></inist:fA14><country>Corée du Sud</country></affiliation></author><author><name sortKey="Choi, Jin Kyu" sort="Choi, Jin Kyu" uniqKey="Choi J" first="Jin-Kyu" last="Choi">Jin-Kyu Choi</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Biotechnology, Yonsei University, 134 Shinchon-dong</s1><s2>Seodaemun-gu, Seoul 120-749</s2><s3>KOR</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ></inist:fA14><country>Corée du Sud</country></affiliation></author><author><name sortKey="Taylor, Deborah R" sort="Taylor, Deborah R" uniqKey="Taylor D" first="Deborah R." last="Taylor">Deborah R. Taylor</name><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Laboratory of Emerging Pathogens, Division of Emerging and Transfusion Transmitted Diseases, Office of Blood Research and Review, Center for Biologics Evaluation and Research, Food and Drug Administration</s1><s2>Bethesda, MD 20892</s2><s3>USA</s3><sZ>3 aut.</sZ></inist:fA14><country>États-Unis</country></affiliation></author><author><name sortKey="Oh, Jong Won" sort="Oh, Jong Won" uniqKey="Oh J" first="Jong-Won" last="Oh">Jong-Won Oh</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Biotechnology, Yonsei University, 134 Shinchon-dong</s1><s2>Seodaemun-gu, Seoul 120-749</s2><s3>KOR</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ></inist:fA14><country>Corée du Sud</country></affiliation><affiliation wicri:level="1"><inist:fA14 i1="03"><s1>Translational Research Center for Protein Function Control, Yonsei University, 134 Shinchon-dong</s1><s2>Seodaemun-gu, Seoul 120-749</s2><s3>KOR</s3><sZ>4 aut.</sZ></inist:fA14><country>Corée du Sud</country></affiliation></author></analytic><series><title level="j" type="main">Archives of virology</title><title level="j" type="abbreviated">Arch. virol.</title><idno type="ISSN">0304-8608</idno><imprint><date when="2012">2012</date></imprint></series></biblStruct></sourceDesc><seriesStmt><title level="j" type="main">Archives of virology</title><title level="j" type="abbreviated">Arch. virol.</title><idno type="ISSN">0304-8608</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>RNA-directed RNA polymerase</term><term>Severe acute respiratory syndrome virus</term></keywords><keywords scheme="Pascal" xml:lang="fr"><term>Virus syndrome respiratoire aigu sévère</term><term>RNA-directed RNA polymerase</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The severe acute respiratory syndrome coronavirus (SARS-CoV) RNA genome is replicated by a virus-encoded RNA replicase, the key component of which is the nonstructural protein 12 (nsp12). In this report, we describe the biochemical properties of a full-length recombinant SARS-CoV nsp12 RNA-dependent RNA polymerase (RdRp) capable of copying viral RNA templates. The purified SARS-CoV nsp12 showed both primer-dependent and primer-independent RNA synthesis activities using homopolymeric RNA templates. The RdRp activity was strictly dependent on Mn<sup>2+</sup>. The nsp12 preferentially copied homopolymeric pyrimidine RNA templates in the absence of an added oligonucleotide primer. It was also able to initiate de novo RNA synthesis from the 3'-ends of both the plus- and minus-strand genome of SARS-CoV, using the 3'-terminal 36- and 37-nt RNA, respectively. 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It was also able to initiate de novo RNA synthesis from the 3'-ends of both the plus- and minus-strand genome of SARS-CoV, using the 3'-terminal 36- and 37-nt RNA, respectively. 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