Maturation mechanism of severe acute respiratory syndrome (SARS) coronavirus 3C-like proteinase.
Identifieur interne : 002500 ( Main/Exploration ); précédent : 002499; suivant : 002501Maturation mechanism of severe acute respiratory syndrome (SARS) coronavirus 3C-like proteinase.
Auteurs : Chunmei Li [République populaire de Chine] ; Yifei Qi ; Xin Teng ; Zongchang Yang ; Ping Wei ; Changsheng Zhang ; Lei Tan ; Lu Zhou ; Ying Liu ; Luhua LaiSource :
- The Journal of biological chemistry [ 1083-351X ] ; 2010.
Descripteurs français
- KwdFr :
- Cysteine endopeptidases (), Cysteine endopeptidases (métabolisme), Inhibiteurs de protéases (pharmacologie), Isatine (pharmacologie), Multimérisation de protéines, Peptides (), Peptides (métabolisme), Polyprotéines (), Polyprotéines (antagonistes et inhibiteurs), Polyprotéines (métabolisme), Protéines virales (), Protéines virales (antagonistes et inhibiteurs), Protéines virales (métabolisme), Protéines à fluorescence verte (métabolisme), Structure quaternaire des protéines, Séquence d'acides aminés, Ultracentrifugation, Virus du SRAS (enzymologie).
- MESH :
- antagonistes et inhibiteurs : Polyprotéines, Protéines virales.
- enzymologie : Virus du SRAS.
- métabolisme : Cysteine endopeptidases, Peptides, Polyprotéines, Protéines virales, Protéines à fluorescence verte.
- pharmacologie : Inhibiteurs de protéases, Isatine.
- Cysteine endopeptidases, Multimérisation de protéines, Peptides, Polyprotéines, Protéines virales, Structure quaternaire des protéines, Séquence d'acides aminés, Ultracentrifugation.
English descriptors
- KwdEn :
- Amino Acid Sequence, Cysteine Endopeptidases (chemistry), Cysteine Endopeptidases (metabolism), Green Fluorescent Proteins (metabolism), Isatin (pharmacology), Peptides (chemistry), Peptides (metabolism), Polyproteins (antagonists & inhibitors), Polyproteins (chemistry), Polyproteins (metabolism), Protease Inhibitors (pharmacology), Protein Multimerization, Protein Structure, Quaternary, SARS Virus (enzymology), Ultracentrifugation, Viral Proteins (antagonists & inhibitors), Viral Proteins (chemistry), Viral Proteins (metabolism).
- MESH :
- chemical , antagonists & inhibitors : Polyproteins, Viral Proteins.
- chemical , chemistry : Cysteine Endopeptidases, Peptides, Polyproteins, Viral Proteins.
- chemical , metabolism : Cysteine Endopeptidases, Green Fluorescent Proteins, Peptides, Polyproteins, Viral Proteins.
- chemical , pharmacology : Isatin, Protease Inhibitors.
- enzymology : SARS Virus.
- Amino Acid Sequence, Protein Multimerization, Protein Structure, Quaternary, Ultracentrifugation.
Abstract
The 3C-like proteinase (3CL(pro)) of the severe acute respiratory syndrome (SARS) coronavirus plays a vital role in virus maturation and is proposed to be a key target for drug design against SARS. Various in vitro studies revealed that only the dimer of the matured 3CL(pro) is active. However, as the internally encoded 3CL(pro) gets matured from the replicase polyprotein by autolytic cleavage at both the N-terminal and the C-terminal flanking sites, it is unclear whether the polyprotein also needs to dimerize first for its autocleavage reaction. We constructed a large protein containing the cyan fluorescent protein (C), the N-terminal flanking substrate peptide of SARS 3CL(pro) (XX), SARS 3CL(pro) (3CLP), and the yellow fluorescent protein (Y) to study the autoprocessing of 3CL(pro) using fluorescence resonance energy transfer. In contrast to the matured 3CL(pro), the polyprotein, as well as the one-step digested product, 3CLP-Y-His, were shown to be monomeric in gel filtration and analytic ultracentrifuge analysis. However, dimers can still be induced and detected when incubating these large proteins with a substrate analog compound in both chemical cross-linking experiments and analytic ultracentrifuge analysis. We also measured enzyme activity under different enzyme concentrations and found a clear tendency of substrate-induced dimer formation. Based on these discoveries, we conclude that substrate-induced dimerization is essential for the activity of SARS-3CL(pro) in the polyprotein, and a modified model for the 3CL(pro) maturation process was proposed. As many viral proteases undergo a similar maturation process, this model might be generally applicable.
DOI: 10.1074/jbc.M109.095851
PubMed: 20489209
Affiliations:
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Le document en format XML
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sort="Qi, Yifei" uniqKey="Qi Y" first="Yifei" last="Qi">Yifei Qi</name></author><author><name sortKey="Teng, Xin" sort="Teng, Xin" uniqKey="Teng X" first="Xin" last="Teng">Xin Teng</name></author><author><name sortKey="Yang, Zongchang" sort="Yang, Zongchang" uniqKey="Yang Z" first="Zongchang" last="Yang">Zongchang Yang</name></author><author><name sortKey="Wei, Ping" sort="Wei, Ping" uniqKey="Wei P" first="Ping" last="Wei">Ping Wei</name></author><author><name sortKey="Zhang, Changsheng" sort="Zhang, Changsheng" uniqKey="Zhang C" first="Changsheng" last="Zhang">Changsheng Zhang</name></author><author><name sortKey="Tan, Lei" sort="Tan, Lei" uniqKey="Tan L" first="Lei" last="Tan">Lei Tan</name></author><author><name sortKey="Zhou, Lu" sort="Zhou, Lu" uniqKey="Zhou L" first="Lu" last="Zhou">Lu Zhou</name></author><author><name sortKey="Liu, Ying" sort="Liu, Ying" uniqKey="Liu Y" first="Ying" last="Liu">Ying Liu</name></author><author><name sortKey="Lai, Luhua" sort="Lai, Luhua" 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type="capitale">Pékin</region></placeName></affiliation></author><author><name sortKey="Qi, Yifei" sort="Qi, Yifei" uniqKey="Qi Y" first="Yifei" last="Qi">Yifei Qi</name></author><author><name sortKey="Teng, Xin" sort="Teng, Xin" uniqKey="Teng X" first="Xin" last="Teng">Xin Teng</name></author><author><name sortKey="Yang, Zongchang" sort="Yang, Zongchang" uniqKey="Yang Z" first="Zongchang" last="Yang">Zongchang Yang</name></author><author><name sortKey="Wei, Ping" sort="Wei, Ping" uniqKey="Wei P" first="Ping" last="Wei">Ping Wei</name></author><author><name sortKey="Zhang, Changsheng" sort="Zhang, Changsheng" uniqKey="Zhang C" first="Changsheng" last="Zhang">Changsheng Zhang</name></author><author><name sortKey="Tan, Lei" sort="Tan, Lei" uniqKey="Tan L" first="Lei" last="Tan">Lei Tan</name></author><author><name sortKey="Zhou, Lu" sort="Zhou, Lu" uniqKey="Zhou L" first="Lu" last="Zhou">Lu Zhou</name></author><author><name sortKey="Liu, Ying" sort="Liu, Ying" uniqKey="Liu Y" first="Ying" last="Liu">Ying Liu</name></author><author><name sortKey="Lai, Luhua" sort="Lai, Luhua" uniqKey="Lai L" first="Luhua" last="Lai">Luhua Lai</name></author></analytic><series><title level="j">The Journal of biological chemistry</title><idno type="eISSN">1083-351X</idno><imprint><date when="2010" type="published">2010</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Amino Acid Sequence</term><term>Cysteine Endopeptidases (chemistry)</term><term>Cysteine Endopeptidases (metabolism)</term><term>Green Fluorescent Proteins (metabolism)</term><term>Isatin (pharmacology)</term><term>Peptides (chemistry)</term><term>Peptides (metabolism)</term><term>Polyproteins (antagonists & inhibitors)</term><term>Polyproteins (chemistry)</term><term>Polyproteins (metabolism)</term><term>Protease Inhibitors (pharmacology)</term><term>Protein Multimerization</term><term>Protein Structure, Quaternary</term><term>SARS Virus (enzymology)</term><term>Ultracentrifugation</term><term>Viral Proteins (antagonists & inhibitors)</term><term>Viral Proteins (chemistry)</term><term>Viral Proteins (metabolism)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Cysteine endopeptidases ()</term><term>Cysteine endopeptidases (métabolisme)</term><term>Inhibiteurs de protéases (pharmacologie)</term><term>Isatine (pharmacologie)</term><term>Multimérisation de protéines</term><term>Peptides ()</term><term>Peptides (métabolisme)</term><term>Polyprotéines ()</term><term>Polyprotéines (antagonistes et inhibiteurs)</term><term>Polyprotéines (métabolisme)</term><term>Protéines virales ()</term><term>Protéines virales (antagonistes et inhibiteurs)</term><term>Protéines virales (métabolisme)</term><term>Protéines à fluorescence verte (métabolisme)</term><term>Structure quaternaire des protéines</term><term>Séquence d'acides aminés</term><term>Ultracentrifugation</term><term>Virus du SRAS (enzymologie)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="antagonists & inhibitors" xml:lang="en"><term>Polyproteins</term><term>Viral Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Cysteine Endopeptidases</term><term>Peptides</term><term>Polyproteins</term><term>Viral Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Cysteine Endopeptidases</term><term>Green Fluorescent Proteins</term><term>Peptides</term><term>Polyproteins</term><term>Viral Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="pharmacology" xml:lang="en"><term>Isatin</term><term>Protease Inhibitors</term></keywords><keywords scheme="MESH" qualifier="antagonistes et inhibiteurs" xml:lang="fr"><term>Polyprotéines</term><term>Protéines virales</term></keywords><keywords scheme="MESH" qualifier="enzymologie" xml:lang="fr"><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="enzymology" xml:lang="en"><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Cysteine endopeptidases</term><term>Peptides</term><term>Polyprotéines</term><term>Protéines virales</term><term>Protéines à fluorescence verte</term></keywords><keywords scheme="MESH" qualifier="pharmacologie" xml:lang="fr"><term>Inhibiteurs de protéases</term><term>Isatine</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Amino Acid Sequence</term><term>Protein Multimerization</term><term>Protein Structure, Quaternary</term><term>Ultracentrifugation</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Cysteine endopeptidases</term><term>Multimérisation de protéines</term><term>Peptides</term><term>Polyprotéines</term><term>Protéines virales</term><term>Structure quaternaire des protéines</term><term>Séquence d'acides aminés</term><term>Ultracentrifugation</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The 3C-like proteinase (3CL(pro)) of the severe acute respiratory syndrome (SARS) coronavirus plays a vital role in virus maturation and is proposed to be a key target for drug design against SARS. Various in vitro studies revealed that only the dimer of the matured 3CL(pro) is active. However, as the internally encoded 3CL(pro) gets matured from the replicase polyprotein by autolytic cleavage at both the N-terminal and the C-terminal flanking sites, it is unclear whether the polyprotein also needs to dimerize first for its autocleavage reaction. We constructed a large protein containing the cyan fluorescent protein (C), the N-terminal flanking substrate peptide of SARS 3CL(pro) (XX), SARS 3CL(pro) (3CLP), and the yellow fluorescent protein (Y) to study the autoprocessing of 3CL(pro) using fluorescence resonance energy transfer. In contrast to the matured 3CL(pro), the polyprotein, as well as the one-step digested product, 3CLP-Y-His, were shown to be monomeric in gel filtration and analytic ultracentrifuge analysis. However, dimers can still be induced and detected when incubating these large proteins with a substrate analog compound in both chemical cross-linking experiments and analytic ultracentrifuge analysis. We also measured enzyme activity under different enzyme concentrations and found a clear tendency of substrate-induced dimer formation. Based on these discoveries, we conclude that substrate-induced dimerization is essential for the activity of SARS-3CL(pro) in the polyprotein, and a modified model for the 3CL(pro) maturation process was proposed. As many viral proteases undergo a similar maturation process, this model might be generally applicable.</div></front></TEI><affiliations><list><country><li>République populaire de Chine</li></country><region><li>Pékin</li></region><settlement><li>Pékin</li></settlement><orgName><li>Université de Pékin</li></orgName></list><tree><noCountry><name sortKey="Lai, Luhua" sort="Lai, Luhua" uniqKey="Lai L" first="Luhua" last="Lai">Luhua Lai</name><name sortKey="Liu, Ying" sort="Liu, Ying" uniqKey="Liu Y" first="Ying" last="Liu">Ying Liu</name><name sortKey="Qi, Yifei" sort="Qi, Yifei" uniqKey="Qi Y" first="Yifei" last="Qi">Yifei Qi</name><name sortKey="Tan, Lei" sort="Tan, Lei" uniqKey="Tan L" first="Lei" last="Tan">Lei Tan</name><name sortKey="Teng, Xin" sort="Teng, Xin" uniqKey="Teng X" first="Xin" last="Teng">Xin Teng</name><name sortKey="Wei, Ping" sort="Wei, Ping" uniqKey="Wei P" first="Ping" last="Wei">Ping Wei</name><name sortKey="Yang, Zongchang" sort="Yang, Zongchang" uniqKey="Yang Z" first="Zongchang" last="Yang">Zongchang Yang</name><name sortKey="Zhang, Changsheng" sort="Zhang, Changsheng" uniqKey="Zhang C" first="Changsheng" last="Zhang">Changsheng Zhang</name><name sortKey="Zhou, Lu" sort="Zhou, Lu" uniqKey="Zhou L" first="Lu" last="Zhou">Lu Zhou</name></noCountry><country name="République populaire de Chine"><noRegion><name sortKey="Li, Chunmei" sort="Li, Chunmei" uniqKey="Li C" first="Chunmei" last="Li">Chunmei Li</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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