Quaternary Structure of the Severe Acute Respiratory Syndrome (SARS) Coronavirus Main Protease

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• 作者：Chi-Yuan Chou ; Hui-Chuan Chang ; Wen-Chi Hsu ; Tien-Zheng Lin ; Chao-Hsiung Lin ; and Gu-Gang Chang
• 刊名：Biochemistry
• 出版年：2004
• 出版时间：November 30, 2004
• 年：2004
• 卷：43
• 期：47
• 页码：14958 - 14970
• 全文大小：1128K
• 年卷期：v.43,no.47(November 30, 2004)
• ISSN：1520-4995

文摘

SARS (severe acute respiratory syndrome) has been one of the most severe viral infectiousdiseases last year and still remains as a highly risky public health problem around the world. Exploringthe types of interactions responsible for structural stabilities of its component protein molecules constitutesone of the approaches to find a destabilization method for the virion particle. In this study, we performeda series of experiments to characterize the quaternary structure of the dimeric coronavirus main protease(M^pro, 3CL^pro). By using the analytical ultracentrifuge, we demonstrated that the dimeric SARS coronavirusmain protease exists as the major form in solution at protein concentration as low as 0.10 mg/mL atneutral pH. The enzyme started to dissociate at acidic and alkali pH values. Ionic strength has profoundeffect on the dimer stability indicating that the major force involved in the subunit association is ionicinteractions. The effect of ionic strength on the protease molecule was reflected by the drastic change ofelectrostatic potential contour of the enzyme in the presence of NaCl. Analysis of the crystal structuresindicated that the interfacial ionic interaction was attributed to the Arg-4···Glu-290 ion pair between thesubunits. Detailed examination of the dimer-monomer equilibrium at different pH values reveals apparentpK_a values of 8.0 ± 0.2 and 5.0 ± 0.1 for the Arg-4 and Glu-290, respectively. Mutation at these twopositions reduces the association affinity between subunits, and the Glu-290 mutants had diminished enzymeactivity. 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.