Effects of metal on the biochemical properties of Helicobacter pylori HypB, a maturation factor of [NiFe]-hydrogenase and urease.
The biosyntheses of the [NiFe]-hydrogenase and urease enzymes in Helicobacter pylori require several accessory proteins for proper construction of the nickel-containing metallocenters. The hydrogenase accessory proteins HypA and HypB, a GTPase, have been implicated in the nickel delivery steps of both enzymes. In this study, the metal-binding properties of H. pylori HypB were characterized, and the effects of metal binding on the biochemical behavior of the protein were examined. The protein can bind stoichiometric amounts of Zn(II) or Ni(II), each with nanomolar affinity. Mutation of Cys106 and His107, which are located between two major GTPase motifs, results in undetectable Ni(II) binding, and the Zn(II) affinity is weakened by 2 orders of magnitude. These two residues are also required for the metal-dependent dimerization observed in the presence of Ni(II) but not Zn(II). The addition of metals to the protein has distinct impacts on GTPase activity, with zinc significantly reducing GTP hydrolysis to below detectable levels and nickel only slightly altering the k(cat) and K(m) of the reaction. The regulation of HypB activities by metal binding may contribute to the maturation of the nickel-containing enzymes.
Department of Chemistry, University of Toronto, 80 St. George Street, Toronto M5S 3H6, Canada.
Andrew M
AM
Sydor
Jenny
J
Liu
Deborah B
DB
Zamble
D019943:Amino Acid Substitution / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000235:genetics; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D016480:Helicobacter pylori / Q000201:enzymology*; D006864:Hydrogenase / Q000378:metabolism*; D007700:Kinetics; D008670:Metals / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D009532:Nickel / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D014510:Urease / Q000378:metabolism*; D015032:Zinc / Q000378:metabolism
D001426:Bacterial Proteins; C089637:HypB protein, bacteria; D008670:Metals; D050505:Mutant Proteins; D009532:Nickel; D006160:Guanosine Triphosphate; C043296:nickel-iron hydrogenase; D006864:Hydrogenase; D014510:Urease; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D015032:Zinc
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