Quaternary Structure of the Severe Acute Respiratory Syndrome (SARS) Coronavirus Main Protease†
Identifieur interne : 000269 ( Istex/Checkpoint ); précédent : 000268; suivant : 000270Quaternary Structure of the Severe Acute Respiratory Syndrome (SARS) Coronavirus Main Protease†
Auteurs : Chi-Yuan Chou [Taïwan] ; Hui-Chuan Chang [Taïwan] ; Wen-Chi Hsu [Taïwan] ; Tien-Zheng Lin [Taïwan] ; Chao-Hsiung Lin [Taïwan] ; Gu-Gang Chang [Taïwan]Source :
- Biochemistry [ 0006-2960 ] ; 2004.
Abstract
SARS (severe acute respiratory syndrome) has been one of the most severe viral infectious diseases last year and still remains as a highly risky public health problem around the world. Exploring the types of interactions responsible for structural stabilities of its component protein molecules constitutes one of the approaches to find a destabilization method for the virion particle. In this study, we performed a series of experiments to characterize the quaternary structure of the dimeric coronavirus main protease (Mpro, 3CLpro). By using the analytical ultracentrifuge, we demonstrated that the dimeric SARS coronavirus main protease exists as the major form in solution at protein concentration as low as 0.10 mg/mL at neutral pH. The enzyme started to dissociate at acidic and alkali pH values. Ionic strength has profound effect on the dimer stability indicating that the major force involved in the subunit association is ionic interactions. The effect of ionic strength on the protease molecule was reflected by the drastic change of electrostatic potential contour of the enzyme in the presence of NaCl. Analysis of the crystal structures indicated that the interfacial ionic interaction was attributed to the Arg-4···Glu-290 ion pair between the subunits. Detailed examination of the dimer−monomer equilibrium at different pH values reveals apparent pKa values of 8.0 ± 0.2 and 5.0 ± 0.1 for the Arg-4 and Glu-290, respectively. Mutation at these two positions reduces the association affinity between subunits, and the Glu-290 mutants had diminished enzyme activity. This information is useful in searching for substances that can intervene in the subunit association, which is attractive as a target to neutralize the virulence of SARS coronavirus.
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DOI: 10.1021/bi0490237
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Coronavirus Main Protease<ref type="bib" target="#bi0490237AF2">†</ref></title><author><name sortKey="Chou, Chi Yuan" sort="Chou, Chi Yuan" uniqKey="Chou C" first="Chi-Yuan" last="Chou">Chi-Yuan Chou</name><affiliation wicri:level="1"><country xml:lang="fr">Taïwan</country><wicri:regionArea>Graduate Institute of Life Sciences, National Defense Medical Center, and Faculty of Life Sciences, Institute of Biochemistry,Structural Biology Program, and Proteome Research Center, National Yang-Ming University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation><affiliation></affiliation></author><author><name sortKey="Chang, Hui Chuan" sort="Chang, Hui Chuan" uniqKey="Chang H" first="Hui-Chuan" last="Chang">Hui-Chuan Chang</name><affiliation wicri:level="1"><country xml:lang="fr">Taïwan</country><wicri:regionArea>Graduate Institute of Life Sciences, National Defense Medical Center, and Faculty of Life Sciences, Institute of Biochemistry,Structural Biology Program, and Proteome Research Center, National Yang-Ming University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation><affiliation></affiliation></author><author><name sortKey="Hsu, Wen Chi" sort="Hsu, Wen Chi" uniqKey="Hsu W" first="Wen-Chi" last="Hsu">Wen-Chi Hsu</name><affiliation wicri:level="1"><country xml:lang="fr">Taïwan</country><wicri:regionArea>Graduate Institute of Life Sciences, National Defense Medical Center, and Faculty of Life Sciences, Institute of Biochemistry,Structural Biology Program, and Proteome Research Center, National Yang-Ming University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation><affiliation></affiliation></author><author><name sortKey="Lin, Tien Zheng" sort="Lin, Tien Zheng" uniqKey="Lin T" first="Tien-Zheng" last="Lin">Tien-Zheng Lin</name><affiliation wicri:level="1"><country xml:lang="fr">Taïwan</country><wicri:regionArea>Graduate Institute of Life Sciences, National Defense Medical Center, and Faculty of Life Sciences, Institute of Biochemistry,Structural Biology Program, and Proteome Research Center, National Yang-Ming University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation><affiliation></affiliation></author><author><name sortKey="Lin, Chao Hsiung" sort="Lin, Chao Hsiung" uniqKey="Lin C" first="Chao-Hsiung" last="Lin">Chao-Hsiung Lin</name><affiliation wicri:level="1"><country xml:lang="fr">Taïwan</country><wicri:regionArea>Graduate Institute of Life Sciences, National Defense Medical Center, and Faculty of Life Sciences, Institute of Biochemistry,Structural Biology Program, and Proteome Research Center, National Yang-Ming University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation><affiliation></affiliation></author><author><name sortKey="Chang, Gu Gang" sort="Chang, Gu Gang" uniqKey="Chang G" first="Gu-Gang" last="Chang">Gu-Gang Chang</name><affiliation wicri:level="1"><country xml:lang="fr">Taïwan</country><wicri:regionArea>Graduate Institute of Life Sciences, National Defense Medical Center, and Faculty of Life Sciences, Institute of Biochemistry,Structural Biology Program, and Proteome Research Center, National Yang-Ming University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation><affiliation></affiliation><affiliation wicri:level="1"><country wicri:rule="url">Taïwan</country></affiliation></author></analytic><monogr></monogr><series><title level="j" type="main">Biochemistry</title><title level="j" type="abbrev">Biochemistry</title><idno type="ISSN">0006-2960</idno><idno type="eISSN">1520-4995</idno><imprint><publisher>American Chemical Society</publisher><date type="e-published" when="2004-10-30">2004</date><date when="2004-11-30">2004</date><biblScope unit="vol">43</biblScope><biblScope unit="issue">47</biblScope><biblScope unit="page" from="14958">14958</biblScope><biblScope unit="page" to="14970">14970</biblScope></imprint><idno type="ISSN">0006-2960</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0006-2960</idno></seriesStmt></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract">SARS (severe acute respiratory syndrome) has been one of the most severe viral infectious diseases last year and still remains as a highly risky public health problem around the world. Exploring the types of interactions responsible for structural stabilities of its component protein molecules constitutes one of the approaches to find a destabilization method for the virion particle. In this study, we performed a series of experiments to characterize the quaternary structure of the dimeric coronavirus main protease (Mpro, 3CLpro). By using the analytical ultracentrifuge, we demonstrated that the dimeric SARS coronavirus main protease exists as the major form in solution at protein concentration as low as 0.10 mg/mL at neutral pH. The enzyme started to dissociate at acidic and alkali pH values. Ionic strength has profound effect on the dimer stability indicating that the major force involved in the subunit association is ionic interactions. The effect of ionic strength on the protease molecule was reflected by the drastic change of electrostatic potential contour of the enzyme in the presence of NaCl. Analysis of the crystal structures indicated that the interfacial ionic interaction was attributed to the Arg-4···Glu-290 ion pair between the subunits. Detailed examination of the dimer−monomer equilibrium at different pH values reveals apparent pKa values of 8.0 ± 0.2 and 5.0 ± 0.1 for the Arg-4 and Glu-290, respectively. Mutation at these two positions reduces the association affinity between subunits, and the Glu-290 mutants had diminished enzyme activity. This information is useful in searching for substances that can intervene in the subunit association, which is attractive as a target to neutralize the virulence of SARS coronavirus.</div></front></TEI><affiliations><list><country><li>Taïwan</li></country></list><tree><country name="Taïwan"><noRegion><name sortKey="Chou, Chi Yuan" sort="Chou, Chi Yuan" uniqKey="Chou C" first="Chi-Yuan" last="Chou">Chi-Yuan Chou</name></noRegion><name sortKey="Chang, Gu Gang" sort="Chang, Gu Gang" uniqKey="Chang G" first="Gu-Gang" last="Chang">Gu-Gang Chang</name><name sortKey="Chang, Gu Gang" sort="Chang, Gu Gang" uniqKey="Chang G" first="Gu-Gang" last="Chang">Gu-Gang Chang</name><name sortKey="Chang, Hui Chuan" sort="Chang, Hui Chuan" uniqKey="Chang H" first="Hui-Chuan" last="Chang">Hui-Chuan Chang</name><name sortKey="Hsu, Wen Chi" sort="Hsu, Wen Chi" uniqKey="Hsu W" first="Wen-Chi" last="Hsu">Wen-Chi Hsu</name><name sortKey="Lin, Chao Hsiung" sort="Lin, Chao Hsiung" uniqKey="Lin C" first="Chao-Hsiung" last="Lin">Chao-Hsiung Lin</name><name sortKey="Lin, Tien Zheng" sort="Lin, Tien Zheng" uniqKey="Lin T" first="Tien-Zheng" last="Lin">Tien-Zheng Lin</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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