Microbial urate catabolism: characterization of HpyO, a non-homologous isofunctional isoform of the flavoprotein urate hydroxylase HpxO
Résumé
In aerobic cells, urate is oxidized to 5-hydroxyisourate
by two distinct enzymes: a coenzyme-independent
urate oxidase (EC 1.7.3.3) found in eukaryotes and
bacteria like Bacillus subtilis and a prokaryotic flavoprotein
urate hydroxylase (HpxO) originally found in
some Klebsiella species. More cases of analogous or
non-homologous isofunctional enzymes (NISE) for
urate catabolism have been hypothesized by inspecting
bacterial genomes. Here, we used a functional
complementation approach in which a candidate gene
for urate oxidation is integrated by homologous
recombination in the Acinetobacter baylyi ADP1
genome at the locus of its original hpxO gene. Catabolism
of urate was restored in A. baylyi ADP1 expressing
a FAD-dependent protein from Xanthomonas
campestris, representing a new urate hydroxylase
family that we called HpyO. This enzyme was kinetically
characterized and compared with other HpxO
enzymes. In contrast to the latter, HpyO is a typical
Michaelian enzyme. This work provides the first
experimental evidences for the function of HpyO in
bacterial urate catabolism and establishes it as a NISE
of HpxO.