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dc.contributor.authorGruschow, Sabine
dc.contributor.authorSadler, Joanna
dc.contributor.authorSharrat, Peter
dc.contributor.authorGoss, Rebecca Jane Miriam
dc.identifier.citationGruschow , S , Sadler , J , Sharrat , P & Goss , R J M 2020 , ' Phenylalanine meta -hydroxylase : a single residue mediates mechanistic control of aromatic amino acid hydroxylation ' , ChemBioChem , vol. 21 , no. 3 , pp. 417-422 .
dc.identifier.otherPURE: 260282409
dc.identifier.otherPURE UUID: 5ce9a7a3-b3cf-4ab9-b559-79a178d36994
dc.identifier.otherRIS: urn:96A4F8AC13D74EC6321266599CF94FEA
dc.identifier.otherScopus: 85075424859
dc.identifier.otherWOS: 000497906200001
dc.descriptionThis work was supported by a project grant from the Biotechnology and Biological Sciences Research Council (BBSRC) U. K to R. J. M. G. (BB/I022910/2), and by the European Research Council under the European Union’s Seventh Framework Programme (FP7-3013/ERC grant agreement no 614779 GenoChemetics).en
dc.description.abstractThe rare non-proteinogenic amino acid, meta- L-tyrosine is biosynthetically intriguing. Whilst the biogenesis of tyrosine from phenylalanine is well characterised, the mechanistic basis for meta-hydroxylation is unknown. Herein, we report the analysis of 3-hydroxylase (Phe3H) from Streptomyces coeruleorbidus. Insight from kinetic analyses, of both the wild-type enzyme and key mutants, of the biocatalytic conversion of synthetic isotopically labelled substrates and fluorinated substrate analogues advances understanding of the process by which meta-hydroxylation is mediated, revealing T202 to play an important role. In contrast to the established mechanism of tyrosine biogenesis, which proceeds via NIH shift, our data support direct, enzyme catalysed deprotonation following electrophilic aromatic substitution. We demonstrate that T202 is responsible for this shift in mechanism, with mutation to alanine resulting in a switch to the NIH shift mechanism and loss of regiospecificity. Furthermore, our kinetic parameters for Phe3H show efficient regiospecific generation of meta-L-tyrosine from phenylalanine and demonstrate the enzyme's ability to regiospecifically hydroxylate unnatural fluorinated substrates.
dc.rightsCopyright © 2019 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.en
dc.subjectMeta-tyrosine biosynthesisen
dc.subjectEnzyme mechanismen
dc.subjectNIH shiften
dc.subjectQD Chemistryen
dc.subjectQH301 Biologyen
dc.titlePhenylalanine meta-hydroxylase : a single residue mediates mechanistic control of aromatic amino acid hydroxylationen
dc.typeJournal articleen
dc.contributor.sponsorEuropean Research Councilen
dc.description.versionPublisher PDFen
dc.contributor.institutionUniversity of St Andrews. School of Biologyen
dc.contributor.institutionUniversity of St Andrews. Biomedical Sciences Research Complexen
dc.contributor.institutionUniversity of St Andrews. School of Chemistryen
dc.contributor.institutionUniversity of St Andrews. EaSTCHEMen
dc.description.statusPeer revieweden

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