Enzymatic activity characterization of SARS coronavirus 3C-like protease by fluorescence resonance energy transfer technique.
Identifieur interne : 002930 ( PubMed/Curation ); précédent : 002929; suivant : 002931Enzymatic activity characterization of SARS coronavirus 3C-like protease by fluorescence resonance energy transfer technique.
Auteurs : Shuai Chen [République populaire de Chine] ; Li-Li Chen ; Hai-Bin Luo ; Tao Sun ; Jing Chen ; Fei Ye ; Jian-Hua Cai ; Jing-Kang Shen ; Xu Shen ; Hua-Liang JiangSource :
- Acta pharmacologica Sinica [ 1671-4083 ] ; 2005.
Descripteurs français
- KwdFr :
- Alanine (génétique), Cinétique, Concentration en ions d'hydrogène, Cysteine endopeptidases, Cystéine (génétique), Endopeptidases (), Endopeptidases (génétique), Histidine (génétique), Mutagenèse dirigée, Méthionine (génétique), Protéines virales (), Protéines virales (génétique), Spécificité du substrat (génétique), Substitution d'acide aminé (génétique), Température, Transfert d'énergie par résonance de fluorescence (), Virus du SRAS (enzymologie), Virus du SRAS (génétique).
- MESH :
- enzymologie : Virus du SRAS.
- génétique : Alanine, Cystéine, Endopeptidases, Histidine, Méthionine, Protéines virales, Spécificité du substrat, Substitution d'acide aminé, Virus du SRAS.
- Cinétique, Concentration en ions d'hydrogène, Cysteine endopeptidases, Endopeptidases, Mutagenèse dirigée, Protéines virales, Température, Transfert d'énergie par résonance de fluorescence.
English descriptors
- KwdEn :
- Alanine (genetics), Amino Acid Substitution (genetics), Cysteine (genetics), Cysteine Endopeptidases, Endopeptidases (chemistry), Endopeptidases (genetics), Fluorescence Resonance Energy Transfer (methods), Histidine (genetics), Hydrogen-Ion Concentration, Kinetics, Methionine (genetics), Mutagenesis, Site-Directed, SARS Virus (enzymology), SARS Virus (genetics), Substrate Specificity (genetics), Temperature, Viral Proteins (chemistry), Viral Proteins (genetics).
- MESH :
- chemical , chemistry : Endopeptidases, Viral Proteins.
- chemical , genetics : Alanine, Cysteine, Endopeptidases, Histidine, Methionine, Viral Proteins.
- enzymology : SARS Virus.
- genetics : Amino Acid Substitution, SARS Virus, Substrate Specificity.
- methods : Fluorescence Resonance Energy Transfer.
- chemical : Cysteine Endopeptidases, Hydrogen-Ion Concentration, Kinetics, Mutagenesis, Site-Directed, Temperature.
Abstract
To characterize enzymatic activity of severe acute respiratory syndrome (SARS) coronavirus (CoV) 3C-like protease (3CL(pro)) and its four site-directed mutants.
DOI: 10.1111/j.1745-7254.2005.00010.x
PubMed: 15659121
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pubmed:15659121Le document en format XML
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<front><div type="abstract" xml:lang="en">To characterize enzymatic activity of severe acute respiratory syndrome (SARS) coronavirus (CoV) 3C-like protease (3CL(pro)) and its four site-directed mutants.</div>
</front>
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<DateCompleted><Year>2005</Year>
<Month>06</Month>
<Day>16</Day>
</DateCompleted>
<DateRevised><Year>2020</Year>
<Month>03</Month>
<Day>25</Day>
</DateRevised>
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<JournalIssue CitedMedium="Print"><Volume>26</Volume>
<Issue>1</Issue>
<PubDate><Year>2005</Year>
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<Title>Acta pharmacologica Sinica</Title>
<ISOAbbreviation>Acta Pharmacol. Sin.</ISOAbbreviation>
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<ArticleTitle>Enzymatic activity characterization of SARS coronavirus 3C-like protease by fluorescence resonance energy transfer technique.</ArticleTitle>
<Pagination><MedlinePgn>99-106</MedlinePgn>
</Pagination>
<Abstract><AbstractText Label="AIM" NlmCategory="OBJECTIVE">To characterize enzymatic activity of severe acute respiratory syndrome (SARS) coronavirus (CoV) 3C-like protease (3CL(pro)) and its four site-directed mutants.</AbstractText>
<AbstractText Label="METHODS" NlmCategory="METHODS">Based on the fluorescence resonance energy transfer (FRET) principle using 5-[(2'-aminoethyl)-amino] naphthelenesulfonic acid (EDANS) and 4-[[4-(dimethylamino) phenyl] azo] benzoic acid (Dabcyl) as the energy transfer pair, one fluorogenic substrate was designed for the evaluation of SARS-CoV 3CL(pro) proteolytic activity.</AbstractText>
<AbstractText Label="RESULTS" NlmCategory="RESULTS">The kinetic parameters of the fluorogenic substrate have been determined as Km=404 micromol.L(-1), kcat=1.08 min(-1), and kcat/Km=2.7 mmol(-1).L.min(-1). SARS-CoV 3CL(pro) showed substantial pH and temperature-triggered activity switches, and site-directed mutagenesis analysis of SARS-CoV 3CL(pro) revealed that substitutions of His41, Cys145, and His163 resulted in complete loss of enzymatic activity, while replacement of Met162 with Ala caused strongly increased activity.</AbstractText>
<AbstractText Label="CONCLUSION" NlmCategory="CONCLUSIONS">This present work has provided valuable information for understanding the catalytic mechanism of SARS-CoV 3CL(pro). This FRET-based assay might supply an ideal approach for the exploration SARS-CoV 3CL(pro) putative inhibitors.</AbstractText>
</Abstract>
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