Protein Hpn: cloning and characterization of a histidine-rich metal-binding polypeptide in Helicobacter pylori and Helicobacter mustelae

Abstract

Helicobacter pylori is a human gastrointestinal pathogen involved in gastritis, duodenal ulcers, and gastric neoplasia. This microorganism produces large amounts of a urease which, like all known ureases, has nickel in the active site. We have identified a protein in clinical isolates of H. pylori and an identical protein in the ferret pathogen Helicobacter mustelae that strongly binds Ni 2؉ and Zn 2؉. This protein has been named Hpn to emphasize its origins in H. pylori and its affinity for nickel. The encoding hpn gene, cloned and expressed in Escherichia coli ER1793, has an open reading frame (180 bp) that specifies a protein with a calculated molecular mass of 7,077 Da and with the same amino-terminal sequence as that of wild-type Hpn. The deduced sequence of Hpn consists of 60 amino acids, of which 28 (47%) are histidines. The hpn gene does not map with the urease gene cluster on the H. pylori chromosome. An Hpn-negative, isogenic H. pylori strain, generated by hpn gene deletion and grown on blood agar, had the same urease activity that wild-type cells did. Thus, the role of Hpn in helicobacters is unknown.

Role of Hpn and NixA of Helicobacter pylori in Susceptibility and Resistance to Bismuth and Other Metal Ions

“…Hpn accounts for approximately 2% of all protein synthesis by H. pylori and is abundant in the cell cytoplasm. As described in the original report [7], H. pylori Hpn is an unusual protein consisting of 60 amino acids, including 28 histidines and four cysteines. The deduced amino acid sequence of H. pylori , H. mustelae , and H. felis Hpn proteins are identical and have stretches of up to seven consecutive histidines.…”

“…This theory was confirmed for nickel and zinc and, to a lesser extent, for cadmium. Indeed, Hpn was first isolated from sonicates of H. pylori by nickel affinity chromatography [7].…”

“…The function of Hpn has not been established, and deletion mutation of the hpn gene in H. pylori does not reduce the measurable urease activity produced in vitro [7]. As this indicates that urease assembly and activity is not Hpn-dependent, we have been interested in defining the phenotypic characteristics of Hpn-deficient cells to explore the functional role of this protein.…”

Hydrogen and Nickel Metabolism inHelicobacterSpecies

“…26 Hpn, first identified in 1995, was named after the protein; it was identified in H. pylori and had affinity for nickel. 56 Disruption of the hpn gene leads to decreased tolerance to nickel concentrations. 57,58 The H. pylori Hpn protein has been recently biochemically characterized: it is mostly found as a multimer in solution, with each monomer of 7 kDa reversibly binding five nickel ions at pH 7.4; nickel can be released by a decrease in pH or in the presence of nickel-chelating, agents such as ethylenediaminetetraacetic acid.…”

“…While both genes can be found in all sequenced H. pylori strains, sequence analysis did not reveal any homolog in H. hepaticus or H. acinonychis. The presence of Hpn was shown in H. mustelae, 56 but it is not known whether the ferret pathogen also possesses Hpn-like. Perhaps Ni-storage/detoxification is most important in the gastric environment where Ni levels would be expected to fluctuate widely.…”