title: Helicobacter pylori HP1512 is a nickel-responsive NikR-regulated outer membrane protein.
abstract: Helicobacter pylori is dependent upon the production of the highly abundant and active metalloenzyme urease for colonization of the human stomach. Thus, H. pylori has an absolute requirement for the transition metal nickel, a required cofactor for urease. To investigate the contribution of genes that are factors in this process, microarray analysis comparing the transcriptome of wild-type H. pylori 26695 cultured in brucella broth containing fetal calf serum (BBF) alone or supplemented with 100 microM NiCl(2) suggested that HP1512 is repressed in the presence of 100 microM supplemental nickel. When measured by comparative real-time quantitative PCR (qPCR), HP1512 transcription was reduced 43-fold relative to the value for the wild type when cultured in BBF supplemented with 10 microM NiCl(2). When grown in unsupplemented BBF, urease activity of an HP1512::cat mutant was significantly reduced compared to the wild type, 4.9 +/- 0.5 micromol/min/mg of protein (n = 7) and 17.1 +/- 4.9 micromol/min/mg of protein (n = 13), respectively (P < 0.0001). In silico analysis of the HP1511-HP1512 (HP1511-1512) intergenic region identified a putative NikR operator upstream of HP1512. Gel shift analysis with purified recombinant NikR verified nickel-dependent binding of H. pylori NikR to the HP1511-1512 intergenic region. Furthermore, comparative real-time qPCR of four nickel-related genes suggests that mutation of HP1512 results in reduced intracellular nickel concentration relative to wild-type H. pylori 26695. Taken together, these data suggest that HP1512 encodes a NikR-nickel-regulated outer membrane protein.
authors.affiliation: Department of Microbiology and Immunology, 5641 Medical Sciences II, University of Michigan Medical School, 1150 W. Medical Center Drive, Ann Arbor, MI 48109-0620, USA.
authors.forename: Gregg S
authors.initials: GS
authors.lastname: Davis
authors.forename: Erika L
authors.initials: EL
authors.lastname: Flannery
authors.forename: Harry L T
authors.initials: HL
authors.lastname: Mobley
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000235:genetics* / Q000502:physiology; D024202:Electrophoretic Mobility Shift Assay; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D016480:Helicobacter pylori / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D009532:Nickel / Q000378:metabolism* / Q000494:pharmacology; D020411:Oligonucleotide Array Sequence Analysis; D012097:Repressor Proteins / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014510:Urease / Q000032:analysis / Q000235:genetics / Q000378:metabolism*
chemical_list: D001425:Bacterial Outer Membrane Proteins; D012097:Repressor Proteins; D009532:Nickel; D014510:Urease