The NfsB-like nitroreductase RdxA in Helicobacter pylori is a flavin reductase: possible implications for metronidazole resistance.

Abstract

The microaerophilic bacterium Helicobacter pylori infects a large proportion of the global human population and is the leading cause for peptic ulcer disease and gastric cancer. The H. pylori eradication therapy rests on four compounds, including the 5-nitroimidazole metronidazole. Unfortunately, metronidazole resistance in H. pylori is an increasingly pressing and severe problem. In most metronidazole-resistant H. pylori isolates mutations in the rdxA gene can be found. RdxA is an NfsB-like nitroreductase and was reported to reduce metronidazole to toxic intermediates. Since nitroreductases can often also act as flavin reductases, we hypothesized that RdxA could reduce flavins. Indeed, we identified RdxA from H. pylori (Hp RdxA) as a potent flavin reductase with a submicromolar K_m for FMN. RdxA also reduced FAD and riboflavin but was inhibited when these substrates were added in low concentrations. Hp RdxA was also inhibited by most nitroaromatic compounds tested. Surprisingly, the enzyme was most strongly inhibited by metronidazole, and no reduction of metronidazole by RdxA could be observed, neither under aerobic nor anaerobic conditions. In contrast, the RdxA homologue from Campylobacter jejuni, Cj RdxA, proved to be an efficient nitroreductase which reduced all nitroaromatic compounds offered, including metronidazole. Cj RdxA also reduced flavins. Based on our observations, we propose a novel, if tentative model in which Hp RdxA does not render H. pylori susceptible to metronidazole by directly reducing the drug's nitro group, but rather by reducing intracellular iron stocks.