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Rationally guided improvement of NOV1 Dioxygenase for the conversion of lignin-derived isoeugenol to vanillin

2022Journal article Restricted access
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dc.contributor.authorDe Simone, Mario
dc.contributor.authorAlvigini, Laura
dc.contributor.authorAlonso-Cotchico, Lur
dc.contributor.authorBrissos, Vânia
dc.contributor.authorCaroli, Jonatan
dc.contributor.authorLucas, Maria Fátima
dc.contributor.authorMonza, Emanuele
dc.contributor.authorMelo, Eduardo
dc.contributor.authorMattevi, Andrea
dc.contributor.authorMartins, Lígia O.
dc.date.accessioned2022-11-30T09:58:39Z
dc.date.available2022-11-30T09:58:39Z
dc.date.issued2022
dc.description.abstractBiocatalysis is a key tool in both green chemistry and biorefinery fields. NOV1 is a dioxygenase that catalyzes the one-step, coenzyme-free oxidation of isoeugenol into vanillin and holds enormous biotechnological potential for the complete valorization of lignin as a sustainable starting material for biobased chemicals, polymers, and materials. This study integrates computational, kinetic, structural, and biophysical approaches to characterize a new NOV1 variant featuring improved activity and stability compared to those of the wild type. The S283F replacement results in a 2-fold increased turnover rate (kcat) for isoeugenol and a 4-fold higher catalytic efficiency (kcat/Km) for molecular oxygen compared to those of the wild type. Furthermore, the variant exhibits a half-life that is 20-fold higher than that of the wild type, which most likely relates to the enhanced stabilization of the iron cofactor in the active site. Molecular dynamics supports this view, revealing that the S283F replacement decreases the optimal pKa and favors conformations of the iron-coordinating histidines compatible with an increased level of binding to iron. Importantly, whole cells containing the S283F variant catalyze the conversion of <= 100 mM isoeugenol to vanillin, yielding >99% molar conversion yields within 24 h. This integrative strategy provided a new enzyme for biotechnological applications and mechanistic insights that will facilitate the future design of robust and efficient biocatalysts.pt_PT
dc.description.sponsorshipPT ALG-01-0145-FEDER-022121
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.doi10.1021/acs.biochem.2c00168pt_PT
dc.identifier.issn0006-2960
dc.identifier.urihttp://hdl.handle.net/10400.1/18557
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherACS Publicationspt_PT
dc.relationLife Sciences for a Healthy and Sustainable Future
dc.relationCentre of Marine Sciences
dc.relationImprovement of Isoeugenol Monooxygenases for Sustainable Production of Lignin-Derived Vanillin
dc.subjectMonooxygenasept_PT
dc.subjectFluorescencept_PT
dc.subjectDynamicpt_PT
dc.subjectEnzymept_PT
dc.titleRationally guided improvement of NOV1 Dioxygenase for the conversion of lignin-derived isoeugenol to vanillinpt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.awardTitleLife Sciences for a Healthy and Sustainable Future
oaire.awardTitleCentre of Marine Sciences
oaire.awardTitleImprovement of Isoeugenol Monooxygenases for Sustainable Production of Lignin-Derived Vanillin
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/LA%2FP%2F0087%2F2020/PT
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UID%2FMulti%2F04326%2F2019/PT
oaire.awardURIinfo:eu-repo/grantAgreement/FCT//2020.08246.BD/PT
oaire.citation.titleBiochemistrypt_PT
oaire.fundingStream6817 - DCRRNI ID
oaire.fundingStream6817 - DCRRNI ID
person.familyNamePinho Melo
person.givenNameEduardo
person.identifier1443188
person.identifier.ciencia-id3C1C-C10C-1510
person.identifier.orcid0000-0002-0974-8977
person.identifier.scopus-author-id35566177900
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.nameFundação para a Ciência e a Tecnologia
project.funder.nameFundação para a Ciência e a Tecnologia
project.funder.nameFundação para a Ciência e a Tecnologia
rcaap.rightsrestrictedAccesspt_PT
rcaap.typearticlept_PT
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relation.isAuthorOfPublication.latestForDiscovery5fa1895f-5577-4652-961a-886ec9bf41b1
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