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Characterisation of thiamine diphosphate-dependent 4-hydroxybenzoylformate decarboxylase enzymes from Rhodococcus jostii RHA1 and Pseudomonas fluorescens Pf-5 involved in degradation of aryl-C2 lignin degradation fragments
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Wei, Zhen, Wilkinson, Rachael C., Rashid, Goran M. M., Brown, David, Fülöp, Vilmos and Bugg, Tim (2019) Characterisation of thiamine diphosphate-dependent 4-hydroxybenzoylformate decarboxylase enzymes from Rhodococcus jostii RHA1 and Pseudomonas fluorescens Pf-5 involved in degradation of aryl-C2 lignin degradation fragments. Biochemistry, 58 (52). pp. 5281-5293. doi:10.1021/acs.biochem.9b00177 ISSN 0006-2960.
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WRAP-Characterisation-thiamine-diphosphate-dependent-Wei-2019.pdf - Accepted Version - Requires a PDF viewer. Download (1270Kb) | Preview |
Official URL: http://dx.doi.org/10.1021/acs.biochem.9b00177
Abstract
A thiamine diphosphate-dependent enzyme annotated as a benzoylformate decarboxylase is encoded in gene cluster ro02984-ro02986 in Rhodococcus jostii RHA1 previously shown to generate vanillin and 4-hydroxybenzaldehyde from lignin oxidation, and a closely related gene cluster is also found in the genome of Pseudomonas fluorescens Pf-5. Two hypotheses for possible pathways involving a thiamine diphosphate-dependent cleavage, either C-C cleavage of a ketol or diketone aryl C3 substrate, or decarboxylation of an aryl C2 substrate, were investigated by expression and purification of the recombinant enzymes, and expression of dehydrogenase and oxidase enzymes also found in the gene clusters. The ThDP-dependent enzymes showed no activity for cleavage of aryl C3 ketol or diketone substrates, but showed activity for decarboxylation of benzoylformate and 4-hydroxybenzoylformate. A flavin-dependent oxidase encoded by gene ro02984 was found to oxidise either mandelic acid or phenylglyoxal. The crystal structure of the P. fluorescens decarboxylase enzyme was determined at 1.69 Å resolution, showing similarity to known benzoylformate decarboxylase enzymes. The P. fluorescens decarboxylase enzyme showed enhanced carboligase activity between vanillin and acetaldehyde, rationalised by the presence of alanine vs serine at residue 73 in the enzyme active site, which was investigated further by site-directed mutagenesis of this residue. A hypothesis for a pathway for degradation of aryl-C2 fragments arising from oxidative cleavage of phenylcoumaran and diarylpropane structures in lignin is proposed.
Item Type: | Journal Article | ||||||||||||||||||
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Subjects: | Q Science > QP Physiology Q Science > QR Microbiology |
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Divisions: | Faculty of Science, Engineering and Medicine > Science > Chemistry | ||||||||||||||||||
Library of Congress Subject Headings (LCSH): | Rhodococcus, Pseudomonas fluorescens, Enzymes, Lignin | ||||||||||||||||||
Journal or Publication Title: | Biochemistry | ||||||||||||||||||
Publisher: | American Chemical Society | ||||||||||||||||||
ISSN: | 0006-2960 | ||||||||||||||||||
Official Date: | 31 December 2019 | ||||||||||||||||||
Dates: |
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Volume: | 58 | ||||||||||||||||||
Number: | 52 | ||||||||||||||||||
Page Range: | pp. 5281-5293 | ||||||||||||||||||
DOI: | 10.1021/acs.biochem.9b00177 | ||||||||||||||||||
Status: | Peer Reviewed | ||||||||||||||||||
Publication Status: | Published | ||||||||||||||||||
Reuse Statement (publisher, data, author rights): | “This document is the Accepted Manuscript version of a Published Work that appeared in final form in Biochemistry, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see http://pubs.acs.org/page/policy/articlesonrequest/index.html].” | ||||||||||||||||||
Access rights to Published version: | Restricted or Subscription Access | ||||||||||||||||||
Date of first compliant deposit: | 16 April 2019 | ||||||||||||||||||
Date of first compliant Open Access: | 4 April 2020 | ||||||||||||||||||
RIOXX Funder/Project Grant: |
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