Mutagenesis of the Mn2+-binding site of manganese peroxidase affects oxidation of Mn2+ by both compound I and compound II.
Identifieur interne : 000C01 ( Main/Exploration ); précédent : 000C00; suivant : 000C02Mutagenesis of the Mn2+-binding site of manganese peroxidase affects oxidation of Mn2+ by both compound I and compound II.
Auteurs : R E Whitwam [États-Unis] ; K R Brown ; M. Musick ; M J Natan ; M. TienSource :
- Biochemistry [ 0006-2960 ] ; 1997.
Descripteurs français
- KwdFr :
- Basidiomycota (enzymologie), Basidiomycota (génétique), Cinétique (MeSH), Composés du fer III (composition chimique), Composés du fer III (métabolisme), Manganèse (métabolisme), Mutagenèse dirigée (MeSH), Oxydoréduction (MeSH), Peroxidases (composition chimique), Peroxidases (génétique), Peroxidases (métabolisme), Sites de fixation (MeSH), Spécificité du substrat (MeSH), Électrochimie (MeSH).
- MESH :
- composition chimique : Composés du fer III, Peroxidases.
- enzymologie : Basidiomycota.
- génétique : Basidiomycota, Peroxidases.
- métabolisme : Composés du fer III, Manganèse, Peroxidases.
- Cinétique, Mutagenèse dirigée, Oxydoréduction, Sites de fixation, Spécificité du substrat, Électrochimie.
English descriptors
- KwdEn :
- Basidiomycota (enzymology), Basidiomycota (genetics), Binding Sites (MeSH), Electrochemistry (MeSH), Ferric Compounds (chemistry), Ferric Compounds (metabolism), Kinetics (MeSH), Manganese (metabolism), Mutagenesis, Site-Directed (MeSH), Oxidation-Reduction (MeSH), Peroxidases (chemistry), Peroxidases (genetics), Peroxidases (metabolism), Substrate Specificity (MeSH).
- MESH :
- chemical , chemistry : Ferric Compounds, Peroxidases.
- enzymology : Basidiomycota.
- genetics : Basidiomycota, Peroxidases.
- chemical , metabolism : Ferric Compounds, Manganese, Peroxidases.
- Binding Sites, Electrochemistry, Kinetics, Mutagenesis, Site-Directed, Oxidation-Reduction, Substrate Specificity.
Abstract
The present study investigates whether compound I and compound II of manganese peroxidase from the white-rot fungus Phanerochaete chrysosporium utilize the same Mn-binding site for catalysis. Manganese peroxidase was expressed from its cDNA in Escherichia coli and refolded from inclusion bodies to yield fully active enzyme. Three mutants of the enzyme were generated by site-directed mutagenesis. Each of the three amino acid residues proposed to be involved in Mn2+ binding, E35, D179, and E39, was mutated. The acidic side chains of E35 and E39 were shortened by one carbon to the acidic group D, and the acidic side chain of D179 was shortened by one carbon to the alkyl group A. These mutants, E35D, D179A, and E39D, were used to determine whether Mn2+ reacts at the same site with both compound I and compound II of manganese peroxidase and to determine whether phenolic substrates for the enzyme react at this site. Our results conclusively demonstrate that E35 and D179 residues are involved not only in Mn2+ binding but also in electron transfer from Mn2+ to the enzyme for both compound I and compound II. In contrast, E39 is not critically important to either process. None of the three residues is involved in reactions with phenolic substrates or with H2O2.
DOI: 10.1021/bi9708794
PubMed: 9245408
Affiliations:
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Musick</name></author><author><name sortKey="Natan, M J" sort="Natan, M J" uniqKey="Natan M" first="M J" last="Natan">M J Natan</name></author><author><name sortKey="Tien, M" sort="Tien, M" uniqKey="Tien M" first="M" last="Tien">M. 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Musick</name></author><author><name sortKey="Natan, M J" sort="Natan, M J" uniqKey="Natan M" first="M J" last="Natan">M J Natan</name></author><author><name sortKey="Tien, M" sort="Tien, M" uniqKey="Tien M" first="M" last="Tien">M. Tien</name></author></analytic><series><title level="j">Biochemistry</title><idno type="ISSN">0006-2960</idno><imprint><date when="1997" type="published">1997</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Basidiomycota (enzymology)</term><term>Basidiomycota (genetics)</term><term>Binding Sites (MeSH)</term><term>Electrochemistry (MeSH)</term><term>Ferric Compounds (chemistry)</term><term>Ferric Compounds (metabolism)</term><term>Kinetics (MeSH)</term><term>Manganese (metabolism)</term><term>Mutagenesis, Site-Directed (MeSH)</term><term>Oxidation-Reduction (MeSH)</term><term>Peroxidases (chemistry)</term><term>Peroxidases (genetics)</term><term>Peroxidases (metabolism)</term><term>Substrate Specificity (MeSH)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Basidiomycota (enzymologie)</term><term>Basidiomycota (génétique)</term><term>Cinétique (MeSH)</term><term>Composés du fer III (composition chimique)</term><term>Composés du fer III (métabolisme)</term><term>Manganèse (métabolisme)</term><term>Mutagenèse dirigée (MeSH)</term><term>Oxydoréduction (MeSH)</term><term>Peroxidases (composition chimique)</term><term>Peroxidases (génétique)</term><term>Peroxidases (métabolisme)</term><term>Sites de fixation (MeSH)</term><term>Spécificité du substrat (MeSH)</term><term>Électrochimie (MeSH)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Ferric Compounds</term><term>Peroxidases</term></keywords><keywords scheme="MESH" qualifier="composition chimique" xml:lang="fr"><term>Composés du fer III</term><term>Peroxidases</term></keywords><keywords scheme="MESH" qualifier="enzymologie" xml:lang="fr"><term>Basidiomycota</term></keywords><keywords scheme="MESH" qualifier="enzymology" xml:lang="en"><term>Basidiomycota</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Basidiomycota</term><term>Peroxidases</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Basidiomycota</term><term>Peroxidases</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Ferric Compounds</term><term>Manganese</term><term>Peroxidases</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Composés du fer III</term><term>Manganèse</term><term>Peroxidases</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Binding Sites</term><term>Electrochemistry</term><term>Kinetics</term><term>Mutagenesis, Site-Directed</term><term>Oxidation-Reduction</term><term>Substrate Specificity</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Cinétique</term><term>Mutagenèse dirigée</term><term>Oxydoréduction</term><term>Sites de fixation</term><term>Spécificité du substrat</term><term>Électrochimie</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The present study investigates whether compound I and compound II of manganese peroxidase from the white-rot fungus Phanerochaete chrysosporium utilize the same Mn-binding site for catalysis. Manganese peroxidase was expressed from its cDNA in Escherichia coli and refolded from inclusion bodies to yield fully active enzyme. Three mutants of the enzyme were generated by site-directed mutagenesis. Each of the three amino acid residues proposed to be involved in Mn2+ binding, E35, D179, and E39, was mutated. The acidic side chains of E35 and E39 were shortened by one carbon to the acidic group D, and the acidic side chain of D179 was shortened by one carbon to the alkyl group A. These mutants, E35D, D179A, and E39D, were used to determine whether Mn2+ reacts at the same site with both compound I and compound II of manganese peroxidase and to determine whether phenolic substrates for the enzyme react at this site. Our results conclusively demonstrate that E35 and D179 residues are involved not only in Mn2+ binding but also in electron transfer from Mn2+ to the enzyme for both compound I and compound II. In contrast, E39 is not critically important to either process. None of the three residues is involved in reactions with phenolic substrates or with H2O2.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">9245408</PMID><DateCompleted><Year>1997</Year><Month>09</Month><Day>08</Day></DateCompleted><DateRevised><Year>2013</Year><Month>11</Month><Day>21</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Print">0006-2960</ISSN><JournalIssue CitedMedium="Print"><Volume>36</Volume><Issue>32</Issue><PubDate><Year>1997</Year><Month>Aug</Month><Day>12</Day></PubDate></JournalIssue><Title>Biochemistry</Title><ISOAbbreviation>Biochemistry</ISOAbbreviation></Journal><ArticleTitle>Mutagenesis of the Mn2+-binding site of manganese peroxidase affects oxidation of Mn2+ by both compound I and compound II.</ArticleTitle><Pagination><MedlinePgn>9766-73</MedlinePgn></Pagination><Abstract><AbstractText>The present study investigates whether compound I and compound II of manganese peroxidase from the white-rot fungus Phanerochaete chrysosporium utilize the same Mn-binding site for catalysis. Manganese peroxidase was expressed from its cDNA in Escherichia coli and refolded from inclusion bodies to yield fully active enzyme. Three mutants of the enzyme were generated by site-directed mutagenesis. Each of the three amino acid residues proposed to be involved in Mn2+ binding, E35, D179, and E39, was mutated. The acidic side chains of E35 and E39 were shortened by one carbon to the acidic group D, and the acidic side chain of D179 was shortened by one carbon to the alkyl group A. These mutants, E35D, D179A, and E39D, were used to determine whether Mn2+ reacts at the same site with both compound I and compound II of manganese peroxidase and to determine whether phenolic substrates for the enzyme react at this site. Our results conclusively demonstrate that E35 and D179 residues are involved not only in Mn2+ binding but also in electron transfer from Mn2+ to the enzyme for both compound I and compound II. In contrast, E39 is not critically important to either process. None of the three residues is involved in reactions with phenolic substrates or with H2O2.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Whitwam</LastName><ForeName>R E</ForeName><Initials>RE</Initials><AffiliationInfo><Affiliation>Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park 16802, USA.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Brown</LastName><ForeName>K R</ForeName><Initials>KR</Initials></Author><Author ValidYN="Y"><LastName>Musick</LastName><ForeName>M</ForeName><Initials>M</Initials></Author><Author ValidYN="Y"><LastName>Natan</LastName><ForeName>M J</ForeName><Initials>MJ</Initials></Author><Author ValidYN="Y"><LastName>Tien</LastName><ForeName>M</ForeName><Initials>M</Initials></Author></AuthorList><Language>eng</Language><GrantList CompleteYN="Y"><Grant><GrantID>GM55312-01</GrantID><Acronym>GM</Acronym><Agency>NIGMS NIH HHS</Agency><Country>United States</Country></Grant></GrantList><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013486">Research Support, U.S. Gov't, Non-P.H.S.</PublicationType><PublicationType UI="D013487">Research Support, U.S. Gov't, P.H.S.</PublicationType></PublicationTypeList></Article><MedlineJournalInfo><Country>United States</Country><MedlineTA>Biochemistry</MedlineTA><NlmUniqueID>0370623</NlmUniqueID><ISSNLinking>0006-2960</ISSNLinking></MedlineJournalInfo><ChemicalList><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D005290">Ferric Compounds</NameOfSubstance></Chemical><Chemical><RegistryNumber>42Z2K6ZL8P</RegistryNumber><NameOfSubstance UI="D008345">Manganese</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 1.11.1.-</RegistryNumber><NameOfSubstance UI="D010544">Peroxidases</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 1.11.1.13</RegistryNumber><NameOfSubstance UI="C051129">manganese peroxidase</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D001487" MajorTopicYN="N">Basidiomycota</DescriptorName><QualifierName UI="Q000201" MajorTopicYN="N">enzymology</QualifierName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D001665" MajorTopicYN="N">Binding Sites</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D004563" MajorTopicYN="N">Electrochemistry</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D005290" MajorTopicYN="N">Ferric Compounds</DescriptorName><QualifierName UI="Q000737" MajorTopicYN="N">chemistry</QualifierName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D007700" MajorTopicYN="N">Kinetics</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D008345" MajorTopicYN="N">Manganese</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D016297" MajorTopicYN="Y">Mutagenesis, Site-Directed</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D010084" MajorTopicYN="N">Oxidation-Reduction</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D010544" MajorTopicYN="N">Peroxidases</DescriptorName><QualifierName UI="Q000737" MajorTopicYN="N">chemistry</QualifierName><QualifierName UI="Q000235" MajorTopicYN="Y">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D013379" MajorTopicYN="N">Substrate Specificity</DescriptorName></MeshHeading></MeshHeadingList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="pubmed"><Year>1997</Year><Month>8</Month><Day>12</Day></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>1997</Year><Month>8</Month><Day>12</Day><Hour>0</Hour><Minute>1</Minute></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>1997</Year><Month>8</Month><Day>12</Day><Hour>0</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">9245408</ArticleId><ArticleId IdType="doi">10.1021/bi9708794</ArticleId><ArticleId IdType="pii">bi9708794</ArticleId></ArticleIdList></PubmedData></pubmed><affiliations><list><country><li>États-Unis</li></country><region><li>Pennsylvanie</li></region><settlement><li>University Park (Pennsylvanie)</li></settlement><orgName><li>Université d'État de Pennsylvanie</li></orgName></list><tree><noCountry><name sortKey="Brown, K R" sort="Brown, K R" uniqKey="Brown K" first="K R" last="Brown">K R Brown</name><name sortKey="Musick, M" sort="Musick, M" uniqKey="Musick M" first="M" last="Musick">M. Musick</name><name sortKey="Natan, M J" sort="Natan, M J" uniqKey="Natan M" first="M J" last="Natan">M J Natan</name><name sortKey="Tien, M" sort="Tien, M" uniqKey="Tien M" first="M" last="Tien">M. Tien</name></noCountry><country name="États-Unis"><region name="Pennsylvanie"><name sortKey="Whitwam, R E" sort="Whitwam, R E" uniqKey="Whitwam R" first="R E" last="Whitwam">R E Whitwam</name></region></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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