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Engineering A-type Dye-Decolorizing Peroxidases by Modification of a Conserved Glutamate Residue

2024-02-20Artigo científico Acesso aberto
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dc.contributor.authorHermann, Enikö
dc.contributor.authorRodrigues, Carolina F.
dc.contributor.authorMartins, Lígia O.
dc.contributor.authorPeterbauer, Clemens
dc.contributor.authorOostenbrink, Chris
dc.contributor.institutionInstituto de Tecnologia Química e Biológica António Xavier (ITQB)
dc.contributor.pblJohn Wiley & Sons, Ltd.
dc.date.accessioned2024-10-01T22:22:54Z
dc.date.available2024-10-01T22:22:54Z
dc.date.issued2024-02-20
dc.descriptionFunding Information: . Financial support by the Austrian Science Fund (FWF) through the doctoral program Biomolecular Technology of Proteins (BioToP, grant number W1224), the Austrian Federal Ministry for Digital and Economic Affairs, the National Foundation for Research, Technology and Development, the Christian Doppler Research Association, Funda\u00E7\u00E3o para a Ci\u00EAncia e Tecnologia (FCT), Portugal, grants FCT 2022.02027.PTDC, MOSTMICRO\u2010ITQB (UIDB/04612/2020 and UIDP/04612/2020), LS4FUTURE (LA/P/0087/2020) are gratefully acknowledged Publisher Copyright: © 2024 The Authors. ChemBioChem published by Wiley-VCH GmbH.
dc.description.abstractDye-decolorizing peroxidases (DyPs) are recently identified microbial enzymes that have been used in several Biotechnology applications from wastewater treatment to lignin valorization. However, their properties and mechanism of action still have many open questions. Their heme-containing active site is buried by three conserved flexible loops with a putative role in modulating substrate access and enzyme catalysis. Here, we investigated the role of a conserved glutamate residue in stabilizing interactions in loop 2 of A-type DyPs. First, we did site saturation mutagenesis of this residue, replacing it with all possible amino acids in bacterial DyPs from Bacillus subtilis (BsDyP) and from Kitasatospora aureofaciens (KaDyP1), the latter being characterized here for the first time. We screened the resulting libraries of variants for activity towards ABTS and identified variants with increased catalytic efficiency. The selected variants were purified and characterized for activity and stability. We furthermore used Molecular Dynamics simulations to rationalize the increased catalytic efficiency and found that the main reason is the electron channeling becoming easier from surface-exposed tryptophans. Based on our findings, we also propose that this glutamate could work as a pH switch in the wild-type enzyme, preventing intracellular damage.en
dc.description.versionpublishersversion
dc.description.versionpublished
dc.format.extent2870327
dc.identifier.doi10.1002/cbic.202300872
dc.identifier.issn1439-4227
dc.identifier.otherPURE: 90406081
dc.identifier.otherPURE UUID: ad1ec573-4429-4696-936f-702f9c463b26
dc.identifier.otherScopus: 85189611886
dc.identifier.otherPubMed: 38376941
dc.identifier.urihttp://hdl.handle.net/10362/172805
dc.identifier.urlhttps://www.scopus.com/pages/publications/85189611886
dc.language.isoeng
dc.peerreviewedyes
dc.subjectEnzymes
dc.subjectOxidoreductases
dc.subjectProtein engineering
dc.subjectProtein structures
dc.subjectStrucutre-activity relationship
dc.subjectBiochemistry
dc.subjectMolecular Medicine
dc.subjectMolecular Biology
dc.subjectOrganic Chemistry
dc.titleEngineering A-type Dye-Decolorizing Peroxidases by Modification of a Conserved Glutamate Residueen
dc.typejournal article
degois.publication.issue9
degois.publication.titleChembiochem
degois.publication.volume25
dspace.entity.typePublication
rcaap.rightsopenAccess

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