Mutation of Asn28 disrupts the dimerization and enzymatic activity of SARS 3CL(pro) .
Identifieur interne : 002128 ( Ncbi/Checkpoint ); précédent : 002127; suivant : 002129Mutation of Asn28 disrupts the dimerization and enzymatic activity of SARS 3CL(pro) .
Auteurs : Jennifer Barrila [États-Unis] ; Sandra B. Gabelli ; Usman Bacha ; L Mario Amzel ; Ernesto FreireSource :
- Biochemistry [ 1520-4995 ] ; 2010.
Descripteurs français
- KwdFr :
- Activation enzymatique (génétique), Asparagine (), Asparagine (génétique), Catalyse, Clonage moléculaire, Cristallographie aux rayons X, Cysteine endopeptidases (), Cysteine endopeptidases (génétique), Cysteine endopeptidases (métabolisme), Données de séquences moléculaires, Modèles moléculaires, Motifs et domaines d'intéraction protéique (génétique), Multimérisation de protéines (génétique), Mutation (physiologie), Protéines virales (), Protéines virales (génétique), Protéines virales (métabolisme), Similitude de séquences d'acides aminés, Substitution d'acide aminé (génétique), Substitution d'acide aminé (physiologie), Séquence d'acides aminés, Virus du SRAS (génétique), Virus du SRAS (métabolisme).
- MESH :
- génétique : Activation enzymatique, Asparagine, Cysteine endopeptidases, Motifs et domaines d'intéraction protéique, Multimérisation de protéines, Protéines virales, Substitution d'acide aminé, Virus du SRAS.
- métabolisme : Cysteine endopeptidases, Protéines virales, Virus du SRAS.
- physiologie : Mutation, Substitution d'acide aminé.
- Asparagine, Catalyse, Clonage moléculaire, Cristallographie aux rayons X, Cysteine endopeptidases, Données de séquences moléculaires, Modèles moléculaires, Protéines virales, Similitude de séquences d'acides aminés, Séquence d'acides aminés.
English descriptors
- KwdEn :
- Amino Acid Sequence, Amino Acid Substitution (genetics), Amino Acid Substitution (physiology), Asparagine (chemistry), Asparagine (genetics), Catalysis, Cloning, Molecular, Crystallography, X-Ray, Cysteine Endopeptidases (chemistry), Cysteine Endopeptidases (genetics), Cysteine Endopeptidases (metabolism), Enzyme Activation (genetics), Models, Molecular, Molecular Sequence Data, Mutation (physiology), Protein Interaction Domains and Motifs (genetics), Protein Multimerization (genetics), SARS Virus (genetics), SARS Virus (metabolism), Sequence Homology, Amino Acid, Viral Proteins (chemistry), Viral Proteins (genetics), Viral Proteins (metabolism).
- MESH :
- chemical , chemistry : Asparagine, Cysteine Endopeptidases, Viral Proteins.
- genetics : Amino Acid Substitution, Asparagine, Cysteine Endopeptidases, Enzyme Activation, Protein Interaction Domains and Motifs, Protein Multimerization, SARS Virus, Viral Proteins.
- chemical , metabolism : Cysteine Endopeptidases, SARS Virus, Viral Proteins.
- physiology : Amino Acid Substitution, Mutation.
- Amino Acid Sequence, Catalysis, Cloning, Molecular, Crystallography, X-Ray, Models, Molecular, Molecular Sequence Data, Sequence Homology, Amino Acid.
Abstract
Coronaviruses are responsible for a significant proportion of annual respiratory and enteric infections in humans and other mammals. The most prominent of these viruses is the severe acute respiratory syndrome coronavirus (SARS-CoV) which causes acute respiratory and gastrointestinal infection in humans. The coronavirus main protease, 3CL(pro), is a key target for broad-spectrum antiviral development because of its critical role in viral maturation and high degree of structural conservation among coronaviruses. Dimerization is an indispensable requirement for the function of SARS 3CL(pro) and is regulated through mechanisms involving both direct and long-range interactions in the enzyme. While many of the binding interactions at the dimerization interface have been extensively studied, those that are important for long-range control are not well-understood. Characterization of these dimerization mechanisms is important for the structure-based design of new treatments targeting coronavirus-based infections. Here we report that Asn28, a residue 11 A from the closest residue in the opposing monomer, is essential for the enzymatic activity and dimerization of SARS 3CL(pro). Mutation of this residue to alanine almost completely inactivates the enzyme and results in a 19.2-fold decrease in the dimerization K(d). The crystallographic structure of the N28A mutant determined at 2.35 A resolution reveals the critical role of Asn28 in maintaining the structural integrity of the active site and in orienting key residues involved in binding at the dimer interface and substrate catalysis. These findings provide deeper insight into complex mechanisms regulating the activity and dimerization of SARS 3CL(pro).
DOI: 10.1021/bi1002585
PubMed: 20420403
Affiliations:
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pubmed:20420403Le document en format XML
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<term>Amino Acid Substitution (physiology)</term>
<term>Asparagine (chemistry)</term>
<term>Asparagine (genetics)</term>
<term>Catalysis</term>
<term>Cloning, Molecular</term>
<term>Crystallography, X-Ray</term>
<term>Cysteine Endopeptidases (chemistry)</term>
<term>Cysteine Endopeptidases (genetics)</term>
<term>Cysteine Endopeptidases (metabolism)</term>
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<term>Molecular Sequence Data</term>
<term>Mutation (physiology)</term>
<term>Protein Interaction Domains and Motifs (genetics)</term>
<term>Protein Multimerization (genetics)</term>
<term>SARS Virus (genetics)</term>
<term>SARS Virus (metabolism)</term>
<term>Sequence Homology, Amino Acid</term>
<term>Viral Proteins (chemistry)</term>
<term>Viral Proteins (genetics)</term>
<term>Viral Proteins (metabolism)</term>
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<term>Asparagine ()</term>
<term>Asparagine (génétique)</term>
<term>Catalyse</term>
<term>Clonage moléculaire</term>
<term>Cristallographie aux rayons X</term>
<term>Cysteine endopeptidases ()</term>
<term>Cysteine endopeptidases (génétique)</term>
<term>Cysteine endopeptidases (métabolisme)</term>
<term>Données de séquences moléculaires</term>
<term>Modèles moléculaires</term>
<term>Motifs et domaines d'intéraction protéique (génétique)</term>
<term>Multimérisation de protéines (génétique)</term>
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<term>Motifs et domaines d'intéraction protéique</term>
<term>Multimérisation de protéines</term>
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<term>Substitution d'acide aminé</term>
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<term>Models, Molecular</term>
<term>Molecular Sequence Data</term>
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<term>Cristallographie aux rayons X</term>
<term>Cysteine endopeptidases</term>
<term>Données de séquences moléculaires</term>
<term>Modèles moléculaires</term>
<term>Protéines virales</term>
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<front><div type="abstract" xml:lang="en">Coronaviruses are responsible for a significant proportion of annual respiratory and enteric infections in humans and other mammals. The most prominent of these viruses is the severe acute respiratory syndrome coronavirus (SARS-CoV) which causes acute respiratory and gastrointestinal infection in humans. The coronavirus main protease, 3CL(pro), is a key target for broad-spectrum antiviral development because of its critical role in viral maturation and high degree of structural conservation among coronaviruses. Dimerization is an indispensable requirement for the function of SARS 3CL(pro) and is regulated through mechanisms involving both direct and long-range interactions in the enzyme. While many of the binding interactions at the dimerization interface have been extensively studied, those that are important for long-range control are not well-understood. Characterization of these dimerization mechanisms is important for the structure-based design of new treatments targeting coronavirus-based infections. Here we report that Asn28, a residue 11 A from the closest residue in the opposing monomer, is essential for the enzymatic activity and dimerization of SARS 3CL(pro). Mutation of this residue to alanine almost completely inactivates the enzyme and results in a 19.2-fold decrease in the dimerization K(d). The crystallographic structure of the N28A mutant determined at 2.35 A resolution reveals the critical role of Asn28 in maintaining the structural integrity of the active site and in orienting key residues involved in binding at the dimer interface and substrate catalysis. These findings provide deeper insight into complex mechanisms regulating the activity and dimerization of SARS 3CL(pro).</div>
</front>
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