Methionine sulphoxide reductase is an important antioxidant enzyme in the gastric pathogen Helicobacter pylori.
The ability of Helicobacter pylori to colonize the stomach requires that it combat oxidative stress responses imposed by the host. The role of methionine sulfoxide reductase (Msr), a methionine repair enzyme, in H. pylori stress resistance was evaluated by a mutant analysis approach. An msr mutant strain lacked immunologically detectable sulphoxide reductase protein and also showed no enzyme activity when provided with oxidized methionines as substrates. The mutant strain showed diminished growth compared to the parent strain in the presence of chemical oxidants, and showed rapid viability loss when exposed to oxidizing conditions. The stress resistance and enzyme activity could be recovered by complementing the mutant with a functional copy of the msr gene. Upon fractionation of parent strain and the complemented mutant cells into membranes and cytoplasmic proteins, most of the immunologically detectable Msr was localized to the membrane, and this fraction contained all of the Msr activity. Qualitative detection of the whole cell protein pattern using 2,4-dinitro phenyl hydrazine (DNPH) showed a far greater number of oxidized protein species in the mutant than in the parent strain when the cells were subjected to oxygen, peroxide or s-nitrosoglutathione (GSNO) induced stress. Importantly, no oxidized proteins were discerned in either strain upon incubation in anaerobic conditions. A mutant strain that synthesized a truncated Msr (corresponding to the MsrA domain) was slightly more resistant to oxidative stress than the msr strain. Mouse colonization studies showed Msr is an important colonization factor, especially for effective longer-term (14 and 21 days) colonization. Complementation of the mutant msr strain by chromosomal insertion of a functional gene restored mouse colonization ability.
Alamuri
Praveen
P
Department of Microbiology, University of Georgia, Athens, GA, USA.
Maier
Robert J
RJ
D000818:Animals; D000975:Antioxidants / Q000378:metabolism*; D004252:DNA Mutational Analysis; D005260:Female; D020868:Gene Silencing; D016480:Helicobacter pylori / Q000187:drug effects / Q000201:enzymology* / Q000254:growth & development / Q000502:physiology; D006801:Humans; D008715:Methionine / Q000737:chemistry / Q000378:metabolism; D051150:Methionine Sulfoxide Reductases; D051379:Mice; D008810:Mice, Inbred C57BL; D016877:Oxidants / Q000494:pharmacology; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D013270:Stomach / Q000382:microbiology*; D013347:Subcellular Fractions / Q000378:metabolism
D000975:Antioxidants; D016877:Oxidants; D008715:Methionine; D010088:Oxidoreductases; D051150:Methionine Sulfoxide Reductases; C027219:methionine sulfoxide reductase