The role of Glu39 in MnII binding and oxidation by manganese peroxidase from Phanerochaete chrysoporium.
Identifieur interne : 000A56 ( Main/Corpus ); précédent : 000A55; suivant : 000A57The role of Glu39 in MnII binding and oxidation by manganese peroxidase from Phanerochaete chrysoporium.
Auteurs : H L Youngs ; M D Sollewijn Gelpke ; D. Li ; M. Sundaramoorthy ; M H GoldSource :
- Biochemistry [ 0006-2960 ] ; 2001.
English descriptors
- KwdEn :
- Amino Acid Substitution (genetics), Aspartic Acid (genetics), Binding Sites (genetics), Computer Simulation (MeSH), Glutamic Acid (metabolism), Kinetics (MeSH), Manganese (metabolism), Models, Molecular (MeSH), Mutagenesis, Site-Directed (MeSH), Oxidation-Reduction (MeSH), Peroxidases (biosynthesis), Peroxidases (genetics), Peroxidases (isolation & purification), Peroxidases (metabolism), Phanerochaete (enzymology), Phanerochaete (genetics), Recombinant Proteins (biosynthesis), Recombinant Proteins (isolation & purification), Recombinant Proteins (metabolism), Sequence Analysis, DNA (MeSH), Spectrophotometry, Ultraviolet (MeSH).
- MESH :
- chemical , biosynthesis : Peroxidases, Recombinant Proteins.
- chemical , genetics : Aspartic Acid, Peroxidases.
- enzymology : Phanerochaete.
- genetics : Amino Acid Substitution, Binding Sites, Phanerochaete.
- chemical , isolation & purification : Peroxidases, Recombinant Proteins.
- chemical , metabolism : Glutamic Acid, Manganese, Peroxidases, Recombinant Proteins.
- Computer Simulation, Kinetics, Models, Molecular, Mutagenesis, Site-Directed, Oxidation-Reduction, Sequence Analysis, DNA, Spectrophotometry, Ultraviolet.
Abstract
Manganese peroxidase (MnP) is a heme-containing enzyme produced by white-rot fungi and is part of the extracellular lignin degrading system in these organisms. MnP is unique among Mn binding enzymes in its ability to bind and oxidize Mn(II) and efficiently release Mn(III). Initial site-directed mutagenesis studies identified the residues E35, E39, and D179 as the Mn binding ligands. However, an E39D variant was recently reported to display wild-type Mn binding and rate of oxidation, calling into question the role of E39 as an Mn ligand. To investigate this hypothesis, we performed computer modeling studies which indicated metal-ligand bond distances in the E39D variant and in an E35D--E39D--D179E triple variant which might allow Mn binding and oxidation. To test the model, we reconstructed the E35D and E39D variants used in the previous study, as well as an E39A single variant and the E35D--E39D--D179E triple variant of MnP isozyme 1 from Phanerochaete chrysosporium. We find that all of the variant proteins are impaired for Mn(II) binding (K(m) increases 20--30-fold) and Mn(II) oxidation (k(cat) decreases 50--400-fold) in both the steady state and the transient state. In particular, mutation of the E39 residue in MnP decreases both Mn binding and oxidation. The catalytic efficiency of the E39A variants decreased approximately 10(4)-fold, while that of the E39D variant decreased approximately 10(3)-fold. Contrary to initial modeling results, the triple variant performed only as well as any of the single Mn ligand variants. Interestingly, the catalytic efficiency of the triple variant decreased only 10(4)-fold, which is approximately 10(2)-fold better than that reported for the E35Q--D179N double variant. These combined studies indicate that precise geometry of the Mn ligands within the Mn binding site of MnP is essential for the efficient binding, oxidation, and release of Mn by this enzyme. The results clearly indicate that E39 is a Mn ligand and that mutation of this ligand decreases both Mn binding and the rate of Mn oxidation.
DOI: 10.1021/bi002104s
PubMed: 11329293
Links to Exploration step
pubmed:11329293Le document en format XML
<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">The role of Glu39 in MnII binding and oxidation by manganese peroxidase from Phanerochaete chrysoporium.</title><author><name sortKey="Youngs, H L" sort="Youngs, H L" uniqKey="Youngs H" first="H L" last="Youngs">H L Youngs</name><affiliation><nlm:affiliation>Department of Biochemistry and Molecular Biology, Oregon Graduate Institute of Science and Technology, Portland, Oregon 97291-1000, USA.</nlm:affiliation></affiliation></author><author><name sortKey="Sollewijn Gelpke, M D" sort="Sollewijn Gelpke, M D" uniqKey="Sollewijn Gelpke M" first="M D" last="Sollewijn Gelpke">M D Sollewijn Gelpke</name></author><author><name sortKey="Li, D" sort="Li, D" uniqKey="Li D" first="D" last="Li">D. Li</name></author><author><name sortKey="Sundaramoorthy, M" sort="Sundaramoorthy, M" uniqKey="Sundaramoorthy M" first="M" last="Sundaramoorthy">M. Sundaramoorthy</name></author><author><name sortKey="Gold, M H" sort="Gold, M H" uniqKey="Gold M" first="M H" last="Gold">M H Gold</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2001">2001</date><idno type="RBID">pubmed:11329293</idno><idno type="pmid">11329293</idno><idno type="doi">10.1021/bi002104s</idno><idno type="wicri:Area/Main/Corpus">000A56</idno><idno type="wicri:explorRef" wicri:stream="Main" wicri:step="Corpus" wicri:corpus="PubMed">000A56</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">The role of Glu39 in MnII binding and oxidation by manganese peroxidase from Phanerochaete chrysoporium.</title><author><name sortKey="Youngs, H L" sort="Youngs, H L" uniqKey="Youngs H" first="H L" last="Youngs">H L Youngs</name><affiliation><nlm:affiliation>Department of Biochemistry and Molecular Biology, Oregon Graduate Institute of Science and Technology, Portland, Oregon 97291-1000, USA.</nlm:affiliation></affiliation></author><author><name sortKey="Sollewijn Gelpke, M D" sort="Sollewijn Gelpke, M D" uniqKey="Sollewijn Gelpke M" first="M D" last="Sollewijn Gelpke">M D Sollewijn Gelpke</name></author><author><name sortKey="Li, D" sort="Li, D" uniqKey="Li D" first="D" last="Li">D. Li</name></author><author><name sortKey="Sundaramoorthy, M" sort="Sundaramoorthy, M" uniqKey="Sundaramoorthy M" first="M" last="Sundaramoorthy">M. Sundaramoorthy</name></author><author><name sortKey="Gold, M H" sort="Gold, M H" uniqKey="Gold M" first="M H" last="Gold">M H Gold</name></author></analytic><series><title level="j">Biochemistry</title><idno type="ISSN">0006-2960</idno><imprint><date when="2001" type="published">2001</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Amino Acid Substitution (genetics)</term><term>Aspartic Acid (genetics)</term><term>Binding Sites (genetics)</term><term>Computer Simulation (MeSH)</term><term>Glutamic Acid (metabolism)</term><term>Kinetics (MeSH)</term><term>Manganese (metabolism)</term><term>Models, Molecular (MeSH)</term><term>Mutagenesis, Site-Directed (MeSH)</term><term>Oxidation-Reduction (MeSH)</term><term>Peroxidases (biosynthesis)</term><term>Peroxidases (genetics)</term><term>Peroxidases (isolation & purification)</term><term>Peroxidases (metabolism)</term><term>Phanerochaete (enzymology)</term><term>Phanerochaete (genetics)</term><term>Recombinant Proteins (biosynthesis)</term><term>Recombinant Proteins (isolation & purification)</term><term>Recombinant Proteins (metabolism)</term><term>Sequence Analysis, DNA (MeSH)</term><term>Spectrophotometry, Ultraviolet (MeSH)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="biosynthesis" xml:lang="en"><term>Peroxidases</term><term>Recombinant Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Aspartic Acid</term><term>Peroxidases</term></keywords><keywords scheme="MESH" qualifier="enzymology" xml:lang="en"><term>Phanerochaete</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Amino Acid Substitution</term><term>Binding Sites</term><term>Phanerochaete</term></keywords><keywords scheme="MESH" type="chemical" qualifier="isolation & purification" xml:lang="en"><term>Peroxidases</term><term>Recombinant Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Glutamic Acid</term><term>Manganese</term><term>Peroxidases</term><term>Recombinant Proteins</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Computer Simulation</term><term>Kinetics</term><term>Models, Molecular</term><term>Mutagenesis, Site-Directed</term><term>Oxidation-Reduction</term><term>Sequence Analysis, DNA</term><term>Spectrophotometry, Ultraviolet</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Manganese peroxidase (MnP) is a heme-containing enzyme produced by white-rot fungi and is part of the extracellular lignin degrading system in these organisms. MnP is unique among Mn binding enzymes in its ability to bind and oxidize Mn(II) and efficiently release Mn(III). Initial site-directed mutagenesis studies identified the residues E35, E39, and D179 as the Mn binding ligands. However, an E39D variant was recently reported to display wild-type Mn binding and rate of oxidation, calling into question the role of E39 as an Mn ligand. To investigate this hypothesis, we performed computer modeling studies which indicated metal-ligand bond distances in the E39D variant and in an E35D--E39D--D179E triple variant which might allow Mn binding and oxidation. To test the model, we reconstructed the E35D and E39D variants used in the previous study, as well as an E39A single variant and the E35D--E39D--D179E triple variant of MnP isozyme 1 from Phanerochaete chrysosporium. We find that all of the variant proteins are impaired for Mn(II) binding (K(m) increases 20--30-fold) and Mn(II) oxidation (k(cat) decreases 50--400-fold) in both the steady state and the transient state. In particular, mutation of the E39 residue in MnP decreases both Mn binding and oxidation. The catalytic efficiency of the E39A variants decreased approximately 10(4)-fold, while that of the E39D variant decreased approximately 10(3)-fold. Contrary to initial modeling results, the triple variant performed only as well as any of the single Mn ligand variants. Interestingly, the catalytic efficiency of the triple variant decreased only 10(4)-fold, which is approximately 10(2)-fold better than that reported for the E35Q--D179N double variant. These combined studies indicate that precise geometry of the Mn ligands within the Mn binding site of MnP is essential for the efficient binding, oxidation, and release of Mn by this enzyme. The results clearly indicate that E39 is a Mn ligand and that mutation of this ligand decreases both Mn binding and the rate of Mn oxidation.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">11329293</PMID><DateCompleted><Year>2001</Year><Month>05</Month><Day>31</Day></DateCompleted><DateRevised><Year>2019</Year><Month>06</Month><Day>13</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Print">0006-2960</ISSN><JournalIssue CitedMedium="Print"><Volume>40</Volume><Issue>7</Issue><PubDate><Year>2001</Year><Month>Feb</Month><Day>20</Day></PubDate></JournalIssue><Title>Biochemistry</Title><ISOAbbreviation>Biochemistry</ISOAbbreviation></Journal><ArticleTitle>The role of Glu39 in MnII binding and oxidation by manganese peroxidase from Phanerochaete chrysoporium.</ArticleTitle><Pagination><MedlinePgn>2243-50</MedlinePgn></Pagination><Abstract><AbstractText>Manganese peroxidase (MnP) is a heme-containing enzyme produced by white-rot fungi and is part of the extracellular lignin degrading system in these organisms. MnP is unique among Mn binding enzymes in its ability to bind and oxidize Mn(II) and efficiently release Mn(III). Initial site-directed mutagenesis studies identified the residues E35, E39, and D179 as the Mn binding ligands. However, an E39D variant was recently reported to display wild-type Mn binding and rate of oxidation, calling into question the role of E39 as an Mn ligand. To investigate this hypothesis, we performed computer modeling studies which indicated metal-ligand bond distances in the E39D variant and in an E35D--E39D--D179E triple variant which might allow Mn binding and oxidation. To test the model, we reconstructed the E35D and E39D variants used in the previous study, as well as an E39A single variant and the E35D--E39D--D179E triple variant of MnP isozyme 1 from Phanerochaete chrysosporium. We find that all of the variant proteins are impaired for Mn(II) binding (K(m) increases 20--30-fold) and Mn(II) oxidation (k(cat) decreases 50--400-fold) in both the steady state and the transient state. In particular, mutation of the E39 residue in MnP decreases both Mn binding and oxidation. The catalytic efficiency of the E39A variants decreased approximately 10(4)-fold, while that of the E39D variant decreased approximately 10(3)-fold. Contrary to initial modeling results, the triple variant performed only as well as any of the single Mn ligand variants. Interestingly, the catalytic efficiency of the triple variant decreased only 10(4)-fold, which is approximately 10(2)-fold better than that reported for the E35Q--D179N double variant. These combined studies indicate that precise geometry of the Mn ligands within the Mn binding site of MnP is essential for the efficient binding, oxidation, and release of Mn by this enzyme. The results clearly indicate that E39 is a Mn ligand and that mutation of this ligand decreases both Mn binding and the rate of Mn oxidation.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Youngs</LastName><ForeName>H L</ForeName><Initials>HL</Initials><AffiliationInfo><Affiliation>Department of Biochemistry and Molecular Biology, Oregon Graduate Institute of Science and Technology, Portland, Oregon 97291-1000, USA.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Sollewijn Gelpke</LastName><ForeName>M D</ForeName><Initials>MD</Initials></Author><Author ValidYN="Y"><LastName>Li</LastName><ForeName>D</ForeName><Initials>D</Initials></Author><Author ValidYN="Y"><LastName>Sundaramoorthy</LastName><ForeName>M</ForeName><Initials>M</Initials></Author><Author ValidYN="Y"><LastName>Gold</LastName><ForeName>M H</ForeName><Initials>MH</Initials></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013486">Research Support, U.S. Gov't, Non-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="D011994">Recombinant Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>30KYC7MIAI</RegistryNumber><NameOfSubstance UI="D001224">Aspartic Acid</NameOfSubstance></Chemical><Chemical><RegistryNumber>3KX376GY7L</RegistryNumber><NameOfSubstance UI="D018698">Glutamic Acid</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="D019943" MajorTopicYN="N">Amino Acid Substitution</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D001224" MajorTopicYN="N">Aspartic Acid</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D001665" MajorTopicYN="N">Binding Sites</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D003198" MajorTopicYN="N">Computer Simulation</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D018698" MajorTopicYN="N">Glutamic Acid</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="Y">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="D008958" MajorTopicYN="N">Models, Molecular</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D016297" MajorTopicYN="N">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="Q000096" MajorTopicYN="N">biosynthesis</QualifierName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000302" MajorTopicYN="N">isolation & purification</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D020075" MajorTopicYN="N">Phanerochaete</DescriptorName><QualifierName UI="Q000201" MajorTopicYN="Y">enzymology</QualifierName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D011994" MajorTopicYN="N">Recombinant Proteins</DescriptorName><QualifierName UI="Q000096" MajorTopicYN="N">biosynthesis</QualifierName><QualifierName UI="Q000302" MajorTopicYN="N">isolation & purification</QualifierName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D017422" MajorTopicYN="N">Sequence Analysis, DNA</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D013056" MajorTopicYN="N">Spectrophotometry, Ultraviolet</DescriptorName></MeshHeading></MeshHeadingList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="pubmed"><Year>2001</Year><Month>5</Month><Day>1</Day><Hour>10</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2001</Year><Month>6</Month><Day>2</Day><Hour>10</Hour><Minute>1</Minute></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>2001</Year><Month>5</Month><Day>1</Day><Hour>10</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">11329293</ArticleId><ArticleId IdType="pii">bi002104s</ArticleId><ArticleId IdType="doi">10.1021/bi002104s</ArticleId></ArticleIdList></PubmedData></pubmed></record>Pour manipuler ce document sous Unix (Dilib)
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