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Department of Molecular Biology & Biotechnology, University of Sheffield, Firth Court, Western Bank, Sheffield S10 2TN, UK
Correspondence
David J. Kelly
d.kelly{at}sheffield.ac.uk
Helicobacter pylori possesses two distinct thioredoxin proteins (Trx1 and Trx2) which may play important roles in the ability of this bacterium to survive oxidative stress. Trx1 has previously been shown to be an electron donor in vitro for alkyl-hydroperoxide reductase (AhpC), one of three members of the peroxiredoxin family of antioxidant peroxidases present in H. pylori. In this study, mutants in the trxA1 and trxA2 genes encoding Trx1 and Trx2, respectively, and in the tpx and bcp genes, which encode the remaining two members of the H. pylori peroxiredoxin family, were constructed in order to determine their roles in resistance to damage by reactive oxygen and nitrogen species. Mutation of trxA1 led to a pronounced increase in sensitivity to oxygen, hydrogen peroxide and the superoxide generator paraquat, as well as to the nitric oxide (NO) releasers sodium nitroprusside (SNP) and S-nitrosoglutathione (GSNO), consistent with an in vivo role for Trx1 as a reductant for AhpC. A trxA2 single mutant grew normally in an atmosphere of 2 % (v/v) O2 but grew very poorly in 10 % (v/v) O2. It showed slight increases in killing by hydrogen peroxide, paraquat, SNP and GSNO compared to the wild-type, but was significantly more sensitive to cumene hydroperoxide in disc-diffusion assays. A trxA1 trxA2 double mutant was very sensitive to all of the oxidative and nitrosative stresses applied. Comparison of the phenotypes of the tpx and bcp mutants showed that Tpx plays a significant role in peroxide and superoxide resistance in H. pylori, while the role of Bcp is minimal. No evidence was obtained for a role for either Tpx or Bcp in resistance to the toxic effects of NO. The results show that a functional thioredoxin system is necessary for both oxidative and nitrosative stress resistance in H. pylori but, surprisingly, is not essential for viability despite the absence of glutathione and a glutaredoxin system in this bacterium.
Abbreviations: GSNO, S-nitrosoglutathione; SNP, sodium nitroprusside
Present address: Microscience Ltd, 545 Eskdale Road, Winnersh Triangle, Wokingham, Berkshire RG41 5TU, UK.
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