Steady-State and Pre-Steady-State Kinetic Evaluation of Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) 3CLpro Cysteine Protease: Development of an Ion-Pair Model for Catalysis
Identifieur interne : 003192 ( Main/Exploration ); précédent : 003191; suivant : 003193Steady-State and Pre-Steady-State Kinetic Evaluation of Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) 3CLpro Cysteine Protease: Development of an Ion-Pair Model for Catalysis
Auteurs : James Solowiej [États-Unis] ; James A. Thomson [États-Unis] ; Kevin Ryan [États-Unis] ; Chun Luo [États-Unis] ; Mingying He [États-Unis] ; Jihong Lou [États-Unis] ; Brion W. Murray [États-Unis, Oman]Source :
- Biochemistry [ 0006-2960 ] ; 2008.
Descripteurs français
- KwdFr :
- 2-Iodo-acétamide (pharmacologie), Acrylates (pharmacocinétique), Activation enzymatique (), Alkylation, Antienzymes (pharmacocinétique), Catalyse, Cinétique, Concentration en ions d'hydrogène, Cysteine endopeptidases (), Cysteine endopeptidases (métabolisme), Cystéine (), Dipeptides (pharmacocinétique), Histidine (), Ions (métabolisme), Modèles biologiques, Protéines virales (), Protéines virales (antagonistes et inhibiteurs), Protéines virales (métabolisme), Solvants (pharmacologie), Stabilité enzymatique (), Transferases (métabolisme), Virus du SRAS (enzymologie), Électricité statique.
- MESH :
- antagonistes et inhibiteurs : Protéines virales.
- enzymologie : Virus du SRAS.
- métabolisme : Cysteine endopeptidases, Ions, Protéines virales, Transferases.
- pharmacocinétique : Acrylates, Antienzymes, Dipeptides.
- pharmacologie : 2-Iodo-acétamide, Solvants.
- Activation enzymatique, Alkylation, Catalyse, Cinétique, Concentration en ions d'hydrogène, Cysteine endopeptidases, Cystéine, Histidine, Modèles biologiques, Protéines virales, Stabilité enzymatique, Électricité statique.
English descriptors
- KwdEn :
- Acrylates (pharmacokinetics), Alkylation, Catalysis, Cysteine (chemistry), Cysteine Endopeptidases (chemistry), Cysteine Endopeptidases (metabolism), Dipeptides (pharmacokinetics), Enzyme Activation (drug effects), Enzyme Inhibitors (pharmacokinetics), Enzyme Stability (drug effects), Histidine (chemistry), Hydrogen-Ion Concentration, Iodoacetamide (pharmacology), Ions (metabolism), Kinetics, Models, Biological, SARS Virus (enzymology), Solvents (pharmacology), Static Electricity, Transferases (metabolism), Viral Proteins (antagonists & inhibitors), Viral Proteins (chemistry), Viral Proteins (metabolism).
- MESH :
- chemical , antagonists & inhibitors : Viral Proteins.
- chemical , chemistry : Cysteine, Cysteine Endopeptidases, Histidine, Viral Proteins.
- chemical , metabolism : Cysteine Endopeptidases, Ions, Transferases, Viral Proteins.
- chemical , pharmacokinetics : Acrylates, Dipeptides, Enzyme Inhibitors.
- drug effects : Enzyme Activation, Enzyme Stability.
- enzymology : SARS Virus.
- chemical , pharmacology : Iodoacetamide, Solvents.
- Alkylation, Catalysis, Hydrogen-Ion Concentration, Kinetics, Models, Biological, Static Electricity.
Abstract
Severe acute respiratory syndrome (SARS) was a worldwide epidemic caused by a coronavirus that has a cysteine protease (3CLpro) essential to its life cycle. Steady-state and pre-steady-state kinetic methods were used with highly active 3CLpro to characterize the reaction mechanism. We show that 3CLpro has mechanistic features common and disparate to the archetypical proteases papain and chymotrypsin. The kinetic mechanism for 3CLpro-mediated ester hydrolysis, including the individual rate constants, is consistent with a simple double displacement mechanism. The pre-steady-state burst rate was independent of ester substrate concentration indicating a high commitment to catalysis. When homologous peptidic amide and ester substrates were compared, a series of interesting observations emerged. Despite a 2000-fold difference in nonenzymatic reactivity, highly related amide and ester substrates were found to have similar kinetic parameters in both the steady-state and pre-steady-state. Steady-state solvent isotope effect (SIE) studies showed an inverse SIE for the amide but not ester substrates. Evaluation of the SIE in the pre-steady-state revealed normal SIEs for both amide and ester burst rates. Proton inventory (PI) studies on amide peptide hydrolysis were consistent with two proton-transfer reactions in the transition state while the ester data was consistent with a single proton-transfer reaction. Finally, the pH-inactivation profile of 3CLpro with iodoacetamide is indicative of an ion-pair mechanism. Taken together, the data are consistent with a 3CLpro mechanism that utilizes an “electrostatic” trigger to initiate the acylation reaction, a cysteine−histidine catalytic dyad ion pair, an enzyme-facilitated release of P1, and a general base-catalyzed deacylation reaction.
Url:
DOI: 10.1021/bi702107v
Affiliations:
Links toward previous steps (curation, corpus...)
- to stream Istex, to step Corpus: 000540
- to stream Istex, to step Curation: 000540
- to stream Istex, to step Checkpoint: 000E66
- to stream PubMed, to step Corpus: 001C23
- to stream PubMed, to step Curation: 001C23
- to stream PubMed, to step Checkpoint: 001A13
- to stream Ncbi, to step Merge: 001B68
- to stream Ncbi, to step Curation: 001B68
- to stream Ncbi, to step Checkpoint: 001B68
- to stream Main, to step Merge: 003294
- to stream Main, to step Curation: 003192
Le document en format XML
<record><TEI wicri:istexFullTextTei="biblStruct"><teiHeader><fileDesc><titleStmt><title>Steady-State and Pre-Steady-State Kinetic Evaluation of Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) 3CLpro Cysteine Protease: Development of an Ion-Pair Model for Catalysis</title><author><name sortKey="Solowiej, James" sort="Solowiej, James" uniqKey="Solowiej J" first="James" last="Solowiej">James Solowiej</name></author><author><name sortKey="Thomson, James A" sort="Thomson, James A" uniqKey="Thomson J" first="James A." last="Thomson">James A. Thomson</name></author><author><name sortKey="Ryan, Kevin" sort="Ryan, Kevin" uniqKey="Ryan K" first="Kevin" last="Ryan">Kevin Ryan</name></author><author><name sortKey="Luo, Chun" sort="Luo, Chun" uniqKey="Luo C" first="Chun" last="Luo">Chun Luo</name></author><author><name sortKey="He, Mingying" sort="He, Mingying" uniqKey="He M" first="Mingying" last="He">Mingying He</name></author><author><name sortKey="Lou, Jihong" sort="Lou, Jihong" uniqKey="Lou J" first="Jihong" last="Lou">Jihong Lou</name></author><author><name sortKey="Murray, Brion W" sort="Murray, Brion W" uniqKey="Murray B" first="Brion W." last="Murray">Brion W. Murray</name></author></titleStmt><publicationStmt><idno type="wicri:source">ISTEX</idno><idno type="RBID">ISTEX:338A9414225E4884248FE4894D84241AEF57F4D3</idno><date when="2008" year="2008">2008</date><idno type="doi">10.1021/bi702107v</idno><idno type="url">https://api.istex.fr/ark:/67375/TPS-6W2RND9C-D/fulltext.pdf</idno><idno type="wicri:Area/Istex/Corpus">000540</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Corpus" wicri:corpus="ISTEX">000540</idno><idno type="wicri:Area/Istex/Curation">000540</idno><idno type="wicri:Area/Istex/Checkpoint">000E66</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Checkpoint">000E66</idno><idno type="wicri:doubleKey">0006-2960:2008:Solowiej J:steady:state:and</idno><idno type="wicri:source">PubMed</idno><idno type="RBID">pubmed:18237196</idno><idno type="wicri:Area/PubMed/Corpus">001C23</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">001C23</idno><idno type="wicri:Area/PubMed/Curation">001C23</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Curation">001C23</idno><idno type="wicri:Area/PubMed/Checkpoint">001A13</idno><idno type="wicri:explorRef" wicri:stream="Checkpoint" wicri:step="PubMed">001A13</idno><idno type="wicri:Area/Ncbi/Merge">001B68</idno><idno type="wicri:Area/Ncbi/Curation">001B68</idno><idno type="wicri:Area/Ncbi/Checkpoint">001B68</idno><idno type="wicri:doubleKey">0006-2960:2008:Solowiej J:steady:state:and</idno><idno type="wicri:Area/Main/Merge">003294</idno><idno type="wicri:Area/Main/Curation">003192</idno><idno type="wicri:Area/Main/Exploration">003192</idno></publicationStmt><sourceDesc><biblStruct><analytic><title level="a" type="main">Steady-State and Pre-Steady-State Kinetic Evaluation of Severe Acute Respiratory
Syndrome Coronavirus (SARS-CoV) 3CL<hi rend="superscript">pro</hi> Cysteine Protease: Development of an
Ion-Pair Model for Catalysis</title><author><name sortKey="Solowiej, James" sort="Solowiej, James" uniqKey="Solowiej J" first="James" last="Solowiej">James Solowiej</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea>Pfizer Global Research and Development, La Jolla, Pfizer Inc., 10777 Science Center Drive, San Diego</wicri:cityArea></affiliation></author><author><name sortKey="Thomson, James A" sort="Thomson, James A" uniqKey="Thomson J" first="James A." last="Thomson">James A. Thomson</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea>Pfizer Global Research and Development, La Jolla, Pfizer Inc., 10777 Science Center Drive, San Diego</wicri:cityArea></affiliation></author><author><name sortKey="Ryan, Kevin" sort="Ryan, Kevin" uniqKey="Ryan K" first="Kevin" last="Ryan">Kevin Ryan</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea>Pfizer Global Research and Development, La Jolla, Pfizer Inc., 10777 Science Center Drive, San Diego</wicri:cityArea></affiliation></author><author><name sortKey="Luo, Chun" sort="Luo, Chun" uniqKey="Luo C" first="Chun" last="Luo">Chun Luo</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea>Pfizer Global Research and Development, La Jolla, Pfizer Inc., 10777 Science Center Drive, San Diego</wicri:cityArea></affiliation><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea> Present address: Accelagen, 11585 Sorrento Valley Road, Suite107, San Diego</wicri:cityArea></affiliation></author><author><name sortKey="He, Mingying" sort="He, Mingying" uniqKey="He M" first="Mingying" last="He">Mingying He</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea>Pfizer Global Research and Development, La Jolla, Pfizer Inc., 10777 Science Center Drive, San Diego</wicri:cityArea></affiliation></author><author><name sortKey="Lou, Jihong" sort="Lou, Jihong" uniqKey="Lou J" first="Jihong" last="Lou">Jihong Lou</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea>Pfizer Global Research and Development, La Jolla, Pfizer Inc., 10777 Science Center Drive, San Diego</wicri:cityArea></affiliation></author><author><name sortKey="Murray, Brion W" sort="Murray, Brion W" uniqKey="Murray B" first="Brion W." last="Murray">Brion W. Murray</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea>Pfizer Global Research and Development, La Jolla, Pfizer Inc., 10777 Science Center Drive, San Diego</wicri:cityArea></affiliation><affiliation wicri:level="1"><country wicri:rule="url">Oman</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">2008</date><date type="published">2008</date><biblScope unit="vol">47</biblScope><biblScope unit="issue">8</biblScope><biblScope unit="page" from="2617">2617</biblScope><biblScope unit="page" to="2630">2630</biblScope></imprint><idno type="ISSN">0006-2960</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0006-2960</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Acrylates (pharmacokinetics)</term><term>Alkylation</term><term>Catalysis</term><term>Cysteine (chemistry)</term><term>Cysteine Endopeptidases (chemistry)</term><term>Cysteine Endopeptidases (metabolism)</term><term>Dipeptides (pharmacokinetics)</term><term>Enzyme Activation (drug effects)</term><term>Enzyme Inhibitors (pharmacokinetics)</term><term>Enzyme Stability (drug effects)</term><term>Histidine (chemistry)</term><term>Hydrogen-Ion Concentration</term><term>Iodoacetamide (pharmacology)</term><term>Ions (metabolism)</term><term>Kinetics</term><term>Models, Biological</term><term>SARS Virus (enzymology)</term><term>Solvents (pharmacology)</term><term>Static Electricity</term><term>Transferases (metabolism)</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>2-Iodo-acétamide (pharmacologie)</term><term>Acrylates (pharmacocinétique)</term><term>Activation enzymatique ()</term><term>Alkylation</term><term>Antienzymes (pharmacocinétique)</term><term>Catalyse</term><term>Cinétique</term><term>Concentration en ions d'hydrogène</term><term>Cysteine endopeptidases ()</term><term>Cysteine endopeptidases (métabolisme)</term><term>Cystéine ()</term><term>Dipeptides (pharmacocinétique)</term><term>Histidine ()</term><term>Ions (métabolisme)</term><term>Modèles biologiques</term><term>Protéines virales ()</term><term>Protéines virales (antagonistes et inhibiteurs)</term><term>Protéines virales (métabolisme)</term><term>Solvants (pharmacologie)</term><term>Stabilité enzymatique ()</term><term>Transferases (métabolisme)</term><term>Virus du SRAS (enzymologie)</term><term>Électricité statique</term></keywords><keywords scheme="MESH" type="chemical" qualifier="antagonists & inhibitors" xml:lang="en"><term>Viral Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Cysteine</term><term>Cysteine Endopeptidases</term><term>Histidine</term><term>Viral Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Cysteine Endopeptidases</term><term>Ions</term><term>Transferases</term><term>Viral Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="pharmacokinetics" xml:lang="en"><term>Acrylates</term><term>Dipeptides</term><term>Enzyme Inhibitors</term></keywords><keywords scheme="MESH" qualifier="antagonistes et inhibiteurs" xml:lang="fr"><term>Protéines virales</term></keywords><keywords scheme="MESH" qualifier="drug effects" xml:lang="en"><term>Enzyme Activation</term><term>Enzyme Stability</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>Ions</term><term>Protéines virales</term><term>Transferases</term></keywords><keywords scheme="MESH" qualifier="pharmacocinétique" xml:lang="fr"><term>Acrylates</term><term>Antienzymes</term><term>Dipeptides</term></keywords><keywords scheme="MESH" qualifier="pharmacologie" xml:lang="fr"><term>2-Iodo-acétamide</term><term>Solvants</term></keywords><keywords scheme="MESH" type="chemical" qualifier="pharmacology" xml:lang="en"><term>Iodoacetamide</term><term>Solvents</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Alkylation</term><term>Catalysis</term><term>Hydrogen-Ion Concentration</term><term>Kinetics</term><term>Models, Biological</term><term>Static Electricity</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Activation enzymatique</term><term>Alkylation</term><term>Catalyse</term><term>Cinétique</term><term>Concentration en ions d'hydrogène</term><term>Cysteine endopeptidases</term><term>Cystéine</term><term>Histidine</term><term>Modèles biologiques</term><term>Protéines virales</term><term>Stabilité enzymatique</term><term>Électricité statique</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract">Severe acute respiratory syndrome (SARS) was a worldwide epidemic caused by a coronavirus that has a cysteine protease (3CLpro) essential to its life cycle. Steady-state and pre-steady-state kinetic methods were used with highly active 3CLpro to characterize the reaction mechanism. We show that 3CLpro has mechanistic features common and disparate to the archetypical proteases papain and chymotrypsin. The kinetic mechanism for 3CLpro-mediated ester hydrolysis, including the individual rate constants, is consistent with a simple double displacement mechanism. The pre-steady-state burst rate was independent of ester substrate concentration indicating a high commitment to catalysis. When homologous peptidic amide and ester substrates were compared, a series of interesting observations emerged. Despite a 2000-fold difference in nonenzymatic reactivity, highly related amide and ester substrates were found to have similar kinetic parameters in both the steady-state and pre-steady-state. Steady-state solvent isotope effect (SIE) studies showed an inverse SIE for the amide but not ester substrates. Evaluation of the SIE in the pre-steady-state revealed normal SIEs for both amide and ester burst rates. Proton inventory (PI) studies on amide peptide hydrolysis were consistent with two proton-transfer reactions in the transition state while the ester data was consistent with a single proton-transfer reaction. Finally, the pH-inactivation profile of 3CLpro with iodoacetamide is indicative of an ion-pair mechanism. Taken together, the data are consistent with a 3CLpro mechanism that utilizes an “electrostatic” trigger to initiate the acylation reaction, a cysteine−histidine catalytic dyad ion pair, an enzyme-facilitated release of P1, and a general base-catalyzed deacylation reaction.</div></front></TEI><affiliations><list><country><li>Oman</li><li>États-Unis</li></country><region><li>Californie</li></region></list><tree><country name="États-Unis"><region name="Californie"><name sortKey="Solowiej, James" sort="Solowiej, James" uniqKey="Solowiej J" first="James" last="Solowiej">James Solowiej</name></region><name sortKey="He, Mingying" sort="He, Mingying" uniqKey="He M" first="Mingying" last="He">Mingying He</name><name sortKey="Lou, Jihong" sort="Lou, Jihong" uniqKey="Lou J" first="Jihong" last="Lou">Jihong Lou</name><name sortKey="Luo, Chun" sort="Luo, Chun" uniqKey="Luo C" first="Chun" last="Luo">Chun Luo</name><name sortKey="Luo, Chun" sort="Luo, Chun" uniqKey="Luo C" first="Chun" last="Luo">Chun Luo</name><name sortKey="Murray, Brion W" sort="Murray, Brion W" uniqKey="Murray B" first="Brion W." last="Murray">Brion W. Murray</name><name sortKey="Ryan, Kevin" sort="Ryan, Kevin" uniqKey="Ryan K" first="Kevin" last="Ryan">Kevin Ryan</name><name sortKey="Thomson, James A" sort="Thomson, James A" uniqKey="Thomson J" first="James A." last="Thomson">James A. Thomson</name></country><country name="Oman"><noRegion><name sortKey="Murray, Brion W" sort="Murray, Brion W" uniqKey="Murray B" first="Brion W." last="Murray">Brion W. Murray</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Sante/explor/SrasV1/Data/Main/Exploration
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 003192 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Main/Exploration/biblio.hfd -nk 003192 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien
|wiki= Sante
|area= SrasV1
|flux= Main
|étape= Exploration
|type= RBID
|clé= ISTEX:338A9414225E4884248FE4894D84241AEF57F4D3
|texte= Steady-State and Pre-Steady-State Kinetic Evaluation of Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) 3CLpro Cysteine Protease: Development of an Ion-Pair Model for Catalysis
}}
| This area was generated with Dilib version V0.6.33. |  |