A putative glutathione-binding site in T4 glutaredoxin investigated by site-directed mutagenesis.
Identifieur interne : 001308 ( Main/Exploration ); précédent : 001307; suivant : 001309A putative glutathione-binding site in T4 glutaredoxin investigated by site-directed mutagenesis.
Auteurs : M. Nikkola ; F K Gleason ; M. Saarinen ; T. Joelson ; O. Björnberg ; H. EklundSource :
- The Journal of biological chemistry [ 0021-9258 ] ; 1991.
Descripteurs français
- KwdFr :
- Alignement de séquences (MeSH), Concentration en ions d'hydrogène (MeSH), Conformation des protéines (MeSH), Disulfures (pharmacologie), Données de séquences moléculaires (MeSH), Escherichia coli (métabolisme), Glutarédoxines (MeSH), Glutathion (métabolisme), Mutagenèse dirigée (MeSH), Oxidoreductases (génétique), Oxidoreductases (métabolisme), Protein-disulfide reductase (glutathione) (MeSH), Protéines (génétique), Protéines (métabolisme), Protéines bactériennes (génétique), Protéines bactériennes (métabolisme), Ribonucleotide reductases (métabolisme), Sites de fixation (MeSH), Spécificité du substrat (MeSH), Séquence d'acides aminés (MeSH).
- MESH :
- génétique : Oxidoreductases, Protéines, Protéines bactériennes.
- métabolisme : Escherichia coli, Glutathion, Oxidoreductases, Protéines, Protéines bactériennes, Ribonucleotide reductases.
- pharmacologie : Disulfures.
- Alignement de séquences, Concentration en ions d'hydrogène, Conformation des protéines, Données de séquences moléculaires, Glutarédoxines, Mutagenèse dirigée, Protein-disulfide reductase (glutathione), Sites de fixation, Spécificité du substrat, Séquence d'acides aminés.
English descriptors
- KwdEn :
- Amino Acid Sequence (MeSH), Bacterial Proteins (genetics), Bacterial Proteins (metabolism), Binding Sites (MeSH), Disulfides (pharmacology), Escherichia coli (metabolism), Glutaredoxins (MeSH), Glutathione (metabolism), Hydrogen-Ion Concentration (MeSH), Molecular Sequence Data (MeSH), Mutagenesis, Site-Directed (MeSH), Oxidoreductases (genetics), Oxidoreductases (metabolism), Protein Conformation (MeSH), Protein Disulfide Reductase (Glutathione) (MeSH), Proteins (genetics), Proteins (metabolism), Ribonucleotide Reductases (metabolism), Sequence Alignment (MeSH), Substrate Specificity (MeSH).
- MESH :
- chemical , genetics : Bacterial Proteins, Oxidoreductases, Proteins.
- chemical , metabolism : Bacterial Proteins, Glutathione, Oxidoreductases, Proteins, Ribonucleotide Reductases.
- chemical , pharmacology : Disulfides.
- metabolism : Escherichia coli.
- Amino Acid Sequence, Binding Sites, Glutaredoxins, Hydrogen-Ion Concentration, Molecular Sequence Data, Mutagenesis, Site-Directed, Protein Conformation, Protein Disulfide Reductase (Glutathione), Sequence Alignment, Substrate Specificity.
Abstract
A glutathione monomer has been docked into the active site cleft of T4 glutaredoxin (previously called T4 thioredoxin) using molecular graphics. The central part of the cleft is formed by the side chain of Tyr-16 on one side and the residues Thr-64, Met-65, and Pro-66 on the other. The entire glutathione molecule fits well into the cleft. A cis-peptide bond between the residues Met-65 and Pro-66 allows glutathione to bind in an anti-parallel fashion to residues 64-66. Hydrogen bonds can be formed between Met-65 and the glutathione cysteine. This binding positions the glutathione sulfur atom ideally for reaction with the glutaredoxin disulfide. In the model, glutathione can form a hydrogen bond to the hydroxyl group of Tyr-16. Charged interactions at opposite ends of the binding cleft are provided by His-12 and Asp-80. The negatively charged alpha-carboxyl group of glutathione may interact with a positive helix dipole of the protein. Fifteen mutant T4 glutaredoxins have been produced and assayed for glutathione binding by determining thioltransferase activity. Mutant proteins with substitutions in the sides of the cleft (Tyr-16, Pro-66) exhibited the most marked decreases in thioltransferase activity. Mutation of His-12 to a serine decreases the catalytic efficiency whereas substitution of Asp-80 by serine increases the catalytic efficiency. A double mutant, D80S;H12S, has much less affinity for glutathione than either single mutant. Substitution of Cys-14 produces an inactive protein, whereas C17S retains some thioltransferase activity.
PubMed: 1874748
Affiliations:
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Björnberg</name></author><author><name sortKey="Eklund, H" sort="Eklund, H" uniqKey="Eklund H" first="H" last="Eklund">H. Eklund</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="1991">1991</date><idno type="RBID">pubmed:1874748</idno><idno type="pmid">1874748</idno><idno type="wicri:Area/Main/Corpus">001296</idno><idno type="wicri:explorRef" wicri:stream="Main" wicri:step="Corpus" wicri:corpus="PubMed">001296</idno><idno type="wicri:Area/Main/Curation">001296</idno><idno type="wicri:explorRef" wicri:stream="Main" wicri:step="Curation">001296</idno><idno type="wicri:Area/Main/Exploration">001296</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">A putative glutathione-binding site in T4 glutaredoxin investigated by site-directed mutagenesis.</title><author><name sortKey="Nikkola, M" sort="Nikkola, M" uniqKey="Nikkola M" first="M" last="Nikkola">M. Nikkola</name><affiliation><nlm:affiliation>Department of Molecular Biology, Swedish University of Agricultural Sciences, Uppsala.</nlm:affiliation><wicri:noCountry code="subField">Uppsala</wicri:noCountry></affiliation></author><author><name sortKey="Gleason, F K" sort="Gleason, F K" uniqKey="Gleason F" first="F K" last="Gleason">F K Gleason</name></author><author><name sortKey="Saarinen, M" sort="Saarinen, M" uniqKey="Saarinen M" first="M" last="Saarinen">M. Saarinen</name></author><author><name sortKey="Joelson, T" sort="Joelson, T" uniqKey="Joelson T" first="T" last="Joelson">T. Joelson</name></author><author><name sortKey="Bjornberg, O" sort="Bjornberg, O" uniqKey="Bjornberg O" first="O" last="Björnberg">O. Björnberg</name></author><author><name sortKey="Eklund, H" sort="Eklund, H" uniqKey="Eklund H" first="H" last="Eklund">H. Eklund</name></author></analytic><series><title level="j">The Journal of biological chemistry</title><idno type="ISSN">0021-9258</idno><imprint><date when="1991" type="published">1991</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Amino Acid Sequence (MeSH)</term><term>Bacterial Proteins (genetics)</term><term>Bacterial Proteins (metabolism)</term><term>Binding Sites (MeSH)</term><term>Disulfides (pharmacology)</term><term>Escherichia coli (metabolism)</term><term>Glutaredoxins (MeSH)</term><term>Glutathione (metabolism)</term><term>Hydrogen-Ion Concentration (MeSH)</term><term>Molecular Sequence Data (MeSH)</term><term>Mutagenesis, Site-Directed (MeSH)</term><term>Oxidoreductases (genetics)</term><term>Oxidoreductases (metabolism)</term><term>Protein Conformation (MeSH)</term><term>Protein Disulfide Reductase (Glutathione) (MeSH)</term><term>Proteins (genetics)</term><term>Proteins (metabolism)</term><term>Ribonucleotide Reductases (metabolism)</term><term>Sequence Alignment (MeSH)</term><term>Substrate Specificity (MeSH)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Alignement de séquences (MeSH)</term><term>Concentration en ions d'hydrogène (MeSH)</term><term>Conformation des protéines (MeSH)</term><term>Disulfures (pharmacologie)</term><term>Données de séquences moléculaires (MeSH)</term><term>Escherichia coli (métabolisme)</term><term>Glutarédoxines (MeSH)</term><term>Glutathion (métabolisme)</term><term>Mutagenèse dirigée (MeSH)</term><term>Oxidoreductases (génétique)</term><term>Oxidoreductases (métabolisme)</term><term>Protein-disulfide reductase (glutathione) (MeSH)</term><term>Protéines (génétique)</term><term>Protéines (métabolisme)</term><term>Protéines bactériennes (génétique)</term><term>Protéines bactériennes (métabolisme)</term><term>Ribonucleotide reductases (métabolisme)</term><term>Sites de fixation (MeSH)</term><term>Spécificité du substrat (MeSH)</term><term>Séquence d'acides aminés (MeSH)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Bacterial Proteins</term><term>Oxidoreductases</term><term>Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Bacterial Proteins</term><term>Glutathione</term><term>Oxidoreductases</term><term>Proteins</term><term>Ribonucleotide Reductases</term></keywords><keywords scheme="MESH" type="chemical" qualifier="pharmacology" xml:lang="en"><term>Disulfides</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Oxidoreductases</term><term>Protéines</term><term>Protéines bactériennes</term></keywords><keywords scheme="MESH" qualifier="metabolism" xml:lang="en"><term>Escherichia coli</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Escherichia coli</term><term>Glutathion</term><term>Oxidoreductases</term><term>Protéines</term><term>Protéines bactériennes</term><term>Ribonucleotide reductases</term></keywords><keywords scheme="MESH" qualifier="pharmacologie" xml:lang="fr"><term>Disulfures</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Amino Acid Sequence</term><term>Binding Sites</term><term>Glutaredoxins</term><term>Hydrogen-Ion Concentration</term><term>Molecular Sequence Data</term><term>Mutagenesis, Site-Directed</term><term>Protein Conformation</term><term>Protein Disulfide Reductase (Glutathione)</term><term>Sequence Alignment</term><term>Substrate Specificity</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Alignement de séquences</term><term>Concentration en ions d'hydrogène</term><term>Conformation des protéines</term><term>Données de séquences moléculaires</term><term>Glutarédoxines</term><term>Mutagenèse dirigée</term><term>Protein-disulfide reductase (glutathione)</term><term>Sites de fixation</term><term>Spécificité du substrat</term><term>Séquence d'acides aminés</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">A glutathione monomer has been docked into the active site cleft of T4 glutaredoxin (previously called T4 thioredoxin) using molecular graphics. The central part of the cleft is formed by the side chain of Tyr-16 on one side and the residues Thr-64, Met-65, and Pro-66 on the other. The entire glutathione molecule fits well into the cleft. A cis-peptide bond between the residues Met-65 and Pro-66 allows glutathione to bind in an anti-parallel fashion to residues 64-66. Hydrogen bonds can be formed between Met-65 and the glutathione cysteine. This binding positions the glutathione sulfur atom ideally for reaction with the glutaredoxin disulfide. In the model, glutathione can form a hydrogen bond to the hydroxyl group of Tyr-16. Charged interactions at opposite ends of the binding cleft are provided by His-12 and Asp-80. The negatively charged alpha-carboxyl group of glutathione may interact with a positive helix dipole of the protein. Fifteen mutant T4 glutaredoxins have been produced and assayed for glutathione binding by determining thioltransferase activity. Mutant proteins with substitutions in the sides of the cleft (Tyr-16, Pro-66) exhibited the most marked decreases in thioltransferase activity. Mutation of His-12 to a serine decreases the catalytic efficiency whereas substitution of Asp-80 by serine increases the catalytic efficiency. A double mutant, D80S;H12S, has much less affinity for glutathione than either single mutant. Substitution of Cys-14 produces an inactive protein, whereas C17S retains some thioltransferase activity.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">1874748</PMID><DateCompleted><Year>1991</Year><Month>09</Month><Day>26</Day></DateCompleted><DateRevised><Year>2013</Year><Month>11</Month><Day>21</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Print">0021-9258</ISSN><JournalIssue CitedMedium="Print"><Volume>266</Volume><Issue>24</Issue><PubDate><Year>1991</Year><Month>Aug</Month><Day>25</Day></PubDate></JournalIssue><Title>The Journal of biological chemistry</Title><ISOAbbreviation>J Biol Chem</ISOAbbreviation></Journal><ArticleTitle>A putative glutathione-binding site in T4 glutaredoxin investigated by site-directed mutagenesis.</ArticleTitle><Pagination><MedlinePgn>16105-12</MedlinePgn></Pagination><Abstract><AbstractText>A glutathione monomer has been docked into the active site cleft of T4 glutaredoxin (previously called T4 thioredoxin) using molecular graphics. The central part of the cleft is formed by the side chain of Tyr-16 on one side and the residues Thr-64, Met-65, and Pro-66 on the other. The entire glutathione molecule fits well into the cleft. A cis-peptide bond between the residues Met-65 and Pro-66 allows glutathione to bind in an anti-parallel fashion to residues 64-66. Hydrogen bonds can be formed between Met-65 and the glutathione cysteine. This binding positions the glutathione sulfur atom ideally for reaction with the glutaredoxin disulfide. In the model, glutathione can form a hydrogen bond to the hydroxyl group of Tyr-16. Charged interactions at opposite ends of the binding cleft are provided by His-12 and Asp-80. The negatively charged alpha-carboxyl group of glutathione may interact with a positive helix dipole of the protein. Fifteen mutant T4 glutaredoxins have been produced and assayed for glutathione binding by determining thioltransferase activity. Mutant proteins with substitutions in the sides of the cleft (Tyr-16, Pro-66) exhibited the most marked decreases in thioltransferase activity. Mutation of His-12 to a serine decreases the catalytic efficiency whereas substitution of Asp-80 by serine increases the catalytic efficiency. A double mutant, D80S;H12S, has much less affinity for glutathione than either single mutant. Substitution of Cys-14 produces an inactive protein, whereas C17S retains some thioltransferase activity.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Nikkola</LastName><ForeName>M</ForeName><Initials>M</Initials><AffiliationInfo><Affiliation>Department of Molecular Biology, Swedish University of Agricultural Sciences, Uppsala.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Gleason</LastName><ForeName>F K</ForeName><Initials>FK</Initials></Author><Author ValidYN="Y"><LastName>Saarinen</LastName><ForeName>M</ForeName><Initials>M</Initials></Author><Author ValidYN="Y"><LastName>Joelson</LastName><ForeName>T</ForeName><Initials>T</Initials></Author><Author ValidYN="Y"><LastName>Björnberg</LastName><ForeName>O</ForeName><Initials>O</Initials></Author><Author ValidYN="Y"><LastName>Eklund</LastName><ForeName>H</ForeName><Initials>H</Initials></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, Non-U.S. Gov't</PublicationType></PublicationTypeList></Article><MedlineJournalInfo><Country>United States</Country><MedlineTA>J Biol Chem</MedlineTA><NlmUniqueID>2985121R</NlmUniqueID><ISSNLinking>0021-9258</ISSNLinking></MedlineJournalInfo><ChemicalList><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D001426">Bacterial Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D004220">Disulfides</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D054477">Glutaredoxins</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D011506">Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 1.-</RegistryNumber><NameOfSubstance UI="D010088">Oxidoreductases</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 1.17.4.-</RegistryNumber><NameOfSubstance UI="D012264">Ribonucleotide Reductases</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 1.8.4.2</RegistryNumber><NameOfSubstance UI="D011490">Protein Disulfide Reductase (Glutathione)</NameOfSubstance></Chemical><Chemical><RegistryNumber>GAN16C9B8O</RegistryNumber><NameOfSubstance UI="D005978">Glutathione</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D000595" MajorTopicYN="N">Amino Acid Sequence</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D001426" MajorTopicYN="N">Bacterial Proteins</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D001665" MajorTopicYN="N">Binding Sites</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D004220" MajorTopicYN="N">Disulfides</DescriptorName><QualifierName UI="Q000494" MajorTopicYN="N">pharmacology</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D004926" MajorTopicYN="N">Escherichia coli</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D054477" MajorTopicYN="N">Glutaredoxins</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D005978" MajorTopicYN="N">Glutathione</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D006863" MajorTopicYN="N">Hydrogen-Ion Concentration</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D008969" MajorTopicYN="N">Molecular Sequence Data</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D016297" MajorTopicYN="N">Mutagenesis, Site-Directed</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D010088" MajorTopicYN="N">Oxidoreductases</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D011487" MajorTopicYN="N">Protein Conformation</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D011490" MajorTopicYN="Y">Protein Disulfide Reductase (Glutathione)</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D011506" MajorTopicYN="N">Proteins</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D012264" MajorTopicYN="N">Ribonucleotide Reductases</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D016415" MajorTopicYN="N">Sequence Alignment</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D013379" MajorTopicYN="N">Substrate Specificity</DescriptorName></MeshHeading></MeshHeadingList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="pubmed"><Year>1991</Year><Month>8</Month><Day>25</Day></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>1991</Year><Month>8</Month><Day>25</Day><Hour>0</Hour><Minute>1</Minute></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>1991</Year><Month>8</Month><Day>25</Day><Hour>0</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">1874748</ArticleId></ArticleIdList></PubmedData></pubmed><affiliations><list></list><tree><noCountry><name sortKey="Bjornberg, O" sort="Bjornberg, O" uniqKey="Bjornberg O" first="O" last="Björnberg">O. Björnberg</name><name sortKey="Eklund, H" sort="Eklund, H" uniqKey="Eklund H" first="H" last="Eklund">H. Eklund</name><name sortKey="Gleason, F K" sort="Gleason, F K" uniqKey="Gleason F" first="F K" last="Gleason">F K Gleason</name><name sortKey="Joelson, T" sort="Joelson, T" uniqKey="Joelson T" first="T" last="Joelson">T. Joelson</name><name sortKey="Nikkola, M" sort="Nikkola, M" uniqKey="Nikkola M" first="M" last="Nikkola">M. Nikkola</name><name sortKey="Saarinen, M" sort="Saarinen, M" uniqKey="Saarinen M" first="M" last="Saarinen">M. Saarinen</name></noCountry></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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