Biochemical and structural explorations of α‐hydroxyacid oxidases reveal a four‐electron oxidative decarboxylation reaction

p‐Hydroxymandelate oxidase (Hmo) is a flavin mononucleotide (FMN)‐dependent enzyme that oxidizes mandelate to benzoylformate. How the FMN‐dependent oxidation is executed by Hmo remains unclear at the molecular level. A continuum of snapshots from crystal structures of Hmo and its mutants in complex...

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Published inActa crystallographica. Section D, Biological crystallography. Vol. 75; no. 8; pp. 733 - 742
Main Authors Yeh, Hsien-Wei, Lin, Kuan-Hung, Lyu, Syue-Yi, Li, Yi-Shan, Huang, Chun-Man, Wang, Yung-Lin, Shih, Hao-Wei, Hsu, Ning-Shian, Wu, Chang-Jer, Li, Tsung-Lin
Format Journal Article
LanguageEnglish
Published 5 Abbey Square, Chester, Cheshire CH1 2HU, England International Union of Crystallography 01.08.2019
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Subjects
Biochemistry
Crystal structure
Crystallography
Decarboxylation
Electrons
Enantiomers
Flavin mononucleotide
hydride transfer
mandelate oxidase
Oxidation
oxidative decarboxylation
Reaction kinetics
Substrate inhibition
α‐hydroxyacids
α-hydroxyacids
mandelate oxidase
flavin mononucleotide
hydride transfer
oxidative decarboxylation
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Summary:p‐Hydroxymandelate oxidase (Hmo) is a flavin mononucleotide (FMN)‐dependent enzyme that oxidizes mandelate to benzoylformate. How the FMN‐dependent oxidation is executed by Hmo remains unclear at the molecular level. A continuum of snapshots from crystal structures of Hmo and its mutants in complex with physiological/nonphysiological substrates, products and inhibitors provides a rationale for its substrate enantioselectivity/promiscuity, its active‐site geometry/reactivity and its direct hydride‐transfer mechanism. A single mutant, Y128F, that extends the two‐electron oxidation reaction to a four‐electron oxidative decarboxylation reaction was unexpectedly observed. Biochemical and structural approaches, including biochemistry, kinetics, stable isotope labeling and X‐ray crystallography, were exploited to reach these conclusions and provide additional insights. Structural and enzymological explorations of α‐hydroxyacid oxidases uncover new flavin mononucleotide‐mediated reactions and intermediates.
ISSN:2059-7983
0907-4449
2059-7983
1399-0047
DOI:10.1107/S2059798319009574