Rapid reaction studies on the chemistry of flavin oxidation in urocanate reductase.
J Biol Chem
; 300(3): 105689, 2024 Mar.
Article
em En
| MEDLINE
| ID: mdl-38280427
ABSTRACT
Urocanate reductase (UrdA) is a bacterial flavin-dependent enzyme that reduces urocanate to imidazole propionate, enabling bacteria to use urocanate as an alternative respiratory electron acceptor. Elevated serum levels of imidazole propionate are associated with the development of type 2 diabetes, and, since UrdA is only present in humans in gut bacteria, this enzyme has emerged as a significant factor linking the health of the gut microbiome and insulin resistance. Here, we investigated the chemistry of flavin oxidation by urocanate in the isolated FAD domain of UrdA (UrdA') using anaerobic stopped-flow experiments. This analysis unveiled the presence of a charge-transfer complex between reduced FAD and urocanate that forms within the dead time of the stopped-flow instrument (â¼1 ms), with flavin oxidation subsequently occurring with a rate constant of â¼60 s-1. The pH dependence of the reaction and analysis of an Arg411Ala mutant of UrdA' are consistent with Arg411 playing a crucial role in catalysis by serving as the active site acid that protonates urocanate during hydride transfer from reduced FAD. Mutational analysis of urocanate-binding residues suggests that the twisted conformation of urocanate imposed by the active site of UrdA' facilitates urocanate reduction. Overall, this study provides valuable insight into the mechanism of urocanate reduction by UrdA.
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Texto completo:
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Base de dados:
MEDLINE
Assunto principal:
Oxirredutases
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Proteínas de Bactérias
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Ácido Urocânico
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Shewanella
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Flavinas
Idioma:
En
Revista:
J Biol Chem
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
Estados Unidos