Relationship between Ni(II) and Zn(II) Coordination and Nucleotide Binding by the Helicobacter pylori [NiFe]-Hydrogenase and Urease Maturation Factor HypB

• Published in: The Journal of biological chemistry 2014-02, Vol.289 (7), p.3828-3841
• Main Authors: Sydor, Andrew M., Lebrette, Hugo, Ariyakumaran, Rishikesh, Cavazza, Christine, Zamble, Deborah B.
• Format: Article
• Language: English
• Subjects: Amino Acid Motifs; Animals; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Cytidine Triphosphate - chemistry; Cytidine Triphosphate - genetics; Cytidine Triphosphate - metabolism; GTP-Binding Proteins - chemistry; GTP-Binding Proteins - genetics; GTP-Binding Proteins - metabolism; Guanosine Diphosphate - chemistry; Guanosine Diphosphate - genetics; Guanosine Diphosphate - metabolism; Helicobacter pylori - enzymology; Helicobacter pylori - genetics; Humans; Hydrogenase; Hydrogenase - chemistry; Hydrogenase - genetics; Hydrogenase - metabolism; Metal Homeostasis; Metalloezyme Biosynthesis; Metalloproteins; Nickel; Nickel - chemistry; Nickel - metabolism; Protein Structure and Folding; Protein Structure, Tertiary; Protein-Metal Ion Interaction; Urease - chemistry; Urease - genetics; Urease - metabolism; Zinc - chemistry; Zinc - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M113.502781

The pathogen Helicobacter pylori requires two nickel-containing enzymes, urease and [NiFe]-hydrogenase, for efficient colonization of the human gastric mucosa. These enzymes possess complex metallocenters that are assembled by teams of proteins in multistep pathways. One essential accessory protein is the GTPase HypB, which is required for Ni(II) delivery to [NiFe]-hydrogenase and participates in urease maturation. Ni(II) or Zn(II) binding to a site embedded in the GTPase domain of HypB modulates the enzymatic activity, suggesting a mechanism of regulation. In this study, biochemical and structural analyses of H. pylori HypB (HpHypB) revealed an intricate link between nucleotide and metal binding. HpHypB nickel coordination, stoichiometry, and affinity were modulated by GTP and GDP, an effect not observed for zinc, and biochemical evidence suggests that His-107 coordination to nickel toggles on and off in a nucleotide-dependent manner. These results are consistent with the crystal structure of HpHypB loaded with Ni(II), GDP, and Pi, which reveals a nickel site distinct from that of zinc-loaded Methanocaldococcus jannaschii HypB as well as subtle changes to the protein structure. Furthermore, Cys-142, a metal ligand from the Switch II GTPase motif, was identified as a key component of the signal transduction between metal binding and the enzymatic activity. Finally, potassium accelerated the enzymatic activity of HpHypB but had no effect on the other biochemical properties of the protein. Altogether, this molecular level information about HpHypB provides insight into its cellular function and illuminates a possible mechanism of metal ion discrimination. Helicobacter pylori HypB (HpHypB) is a metal-regulated GTPase essential for the biosynthesis of [NiFe]-hydrogenase and urease. Nickel binding to HpHypB is altered upon nucleotide binding, an effect not observed with zinc. Metal coordination and the GTPase cycle of HpHypB are intimately linked. These data suggest that HpHypB contributes to metal fidelity in [NiFe]-hydrogenase and urease biosynthesis.