Nickel Metabolism in Helicobacter pylori

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DESCRIPTION (provided by applicant): Helicobacter pyori is a spiral bacterium that colonizes the gastric mucosa of humans, leading to a variety of gastric diseases that include peptic ulcers, chronic gastritis, mucosal-associated lymphomas, and adenocarcinomas of the lower stomach and duodenum. Two nickel-containing enzymes, urease and hydrogenase, are important for the bacterium's mucosal-colonizing abilities. The goal proposed here is to understand the nickel sequestering, storage, metabolizing and metalloregulatory steps involved in the synthesis/maturation of the two nickel-containing enzymes. From studies with other organisms, the sequence-identified accessory proteins (encoded by ure and hyp genes) would be expected to form nickel sequestering and energy utilizing (GTP hydrolyzing) complexes needed for maturation of the two Ni-enzymes. In the presence of nickel, these complexes facilitate mobilization of the metal into the final sink, urease or hydrogenase. From our knowledge of purified nickel binding and GTPase accessory proteins and their interactions, a sequential Ni-mobilizing transfer pathway will be proposed. The hypothesized Ni-transfer steps will be tested by mixing together selected pure accessory proteins. The roles of some specific nickel-binding proteins (predicted from the genome sequence) in nickel sequestering, homeostasis, and regulation will be addressed via targeted mutagenesis. The specific approaches to understand nickel metabolism, homeostasis, and regulation, in H. pylori will include purification of complexes that deliver nickel to the Ni-enzymes, proteomic analysis to identify proteins, and genomic analysis of transcripts that are regulated in response to nickel supplementation. Finally, gene directed mutants in proteins that are anticipated to affect the nickel nutritional status of the bacterium (i.e. nickel homeostasis) will be characterized to understand the roles of three specific (but poorly studied) nickel binding proteins. The role of a DNA binding and nickel-sensing protein (NikR) is of particular interest as it may play a global regulatory role in metal homeostasis, in turn affecting many processes of cell metabolism.