Contribution of the Helicobacter pylori Thiol Peroxidase Bacterioferritin Comigratory Protein to Oxidative Stress Resistance and Host Colonization

Oxidative stress resistance is one of the key properties that enable pathogenic bacteria to survive the effects of the production of reactive oxygen by the host (21). Peroxiredoxins (Prx) are ubiquitous proteins that confer resistance to oxidative stress. They are enzymes lacking prosthetic groups that catalyze the reduction of hydrogen peroxide and organic hydroperoxides (26). Peroxiredoxins can be divided into two subgroups according to the number of conserved cysteines (Cys) within the proteins. Some peroxiredoxins contain only a single essential, N-terminal Cys residue per subunit (1-Cys Prx), and other peroxiredoxins contain an additional conserved Cys residue that links the two subunits via an intersubunit disulfide bond with the N-terminal Cys in the oxidized protein (2-Cys Prx). Helicobacter pylori is a microaerophilic bacterium that causes peptic ulcers and is a risk factor for adenocarcinomas in humans (7). H. pylori contains three members of the Prx family of reductases, with the most important one being AhpC (alkyl hydroperoxide reductase). H. pylori AhpC is a 2-Cys Prx, and it uses reduced thioredoxin (Trx) as the electron donor to reduce hydrogen peroxide and organic hydroperoxides (3). It was also shown to have activity in reducing peroxynitrite (5). H. pylori AhpC plays a very important role in oxidative stress resistance and host colonization (16, 17). In addition to ahpC, two genes, tpx (HP0390) and bcp (HP0136), coding for two thiol peroxidases that belong to a 1-Cys Prx subgroup have been revealed by H. pylori genome sequence annotation (1, 22). Tpx was originally identified as an Escherichia coli homologue of scavengase p20, and a thioredoxin-linked peroxidase activity of Tpx protein was demonstrated (23, 28). An H. pylori tpx mutant strain is more sensitive to killing by peroxide and superoxide than the wild-type strain, and it has reduced ability to colonize mouse stomachs (6, 17). Bacterioferritin comigratory protein (BCP), originally identified in E. coli, was shown to be a new member of 1-Cys Prx. E. coli BCP shows a thioredoxin-dependent thiol peroxidase activity, with linoleic acid hydroperoxide (LOOH) being the preferred substrate over H2O2 (12). An E. coli bcp mutant shows the same level of hypersensitivity to H2O2 and organic peroxides as an ahpC mutant (12). This study was initiated to characterize H. pylori BCP and to investigate the roles of this protein in oxidative stress resistance and in the colonization of the host stomach. Recently, Comtois et al. (6) constructed an H. pylori bcp mutant (in strain 26695) and observed a weak phenotype in vitro; thus, no further investigation was conducted. In the present study, we characterized in more detail the bcp mutants from different strains, particularly in comparison with an ahpC mutant strain. In addition, we purified H. pylori BCP and determined its peroxidase activity, which showed that BCP has a function overlapping that of AhpC. We report that the bcp mutant displays a relatively weak phenotype because of the presence of much more abundant AhpC within H. pylori cells. More importantly, with mouse colonization studies we demonstrated that H. pylori BCP contributes significantly to the bacterium's ability to colonize the host stomach, particularly in longer-term (3-week) colonization assays.