ИСТИНА

We performed heterologous expression of the gene of the hypothetical UrdA (SO_4620) protein from Shewanella oneidensis MR-1 [1]. The recombinant protein was purified and investigated. It was shown that UrdA contains two flavin prosthetic groups: a noncovalently bound FAD and a covalently bound FMN residue. Redox titration of these centers at pH 7.5 showed that the noncovalently bound FAD operates as a two-electron carrier with midpoint potential of –280 mV, while the covalently bound FMN residue capable of two sequential one-electron reduction steps (Em1 = –225 mV and Em2 = –250 mV) with transient formation of the anionic semiquinone. Using structural modeling and comparative genomic analysis we proposed that UrdA catalyses reduction of urocanate in bacterial cells. The following experiments confirmed this prediction and showed that UrdA does catalyse the unidirectional reaction of 2-electron reduction of urocanic acid to deamino-histidine, an activity not reported earlier. UrdA exhibits both high substrate affinity and high turnover rate (Km << 10 mkM, kcat = 360 s-1) and strong specificity in favor of urocanic acid. UrdA homologues are present in various bacterial genera, such as Shewanella, Fusobacterium, and Clostridium, the latter including the human pathogen Clostridium tetani. The UrdA activity in S. oneidensis is induced by its substrate under anaerobic conditions and it enables anaerobic growth with urocanic acid as a sole terminal electron acceptor. The latter capability can provide the cells of UrdA-containing bacteria with a niche where no other bacteria can compete and survive.