Helicobacter pylori hydrogenase accessory protein HypA and urease accessory protein UreG compete with each other for UreE recognition

Background: The gastric pathogen Helicobacter pylori relies on nickel-containing urease and hydrogenase enzymes in order to colonize the host. Incorporation of Ni2+ into urease is essential for the function of the enzyme and requires the action of several accessory proteins, including the hydrogenase accessory proteins HypA and HypB and the urease accessory proteins UreE, UreF, UreG and UreH. Methods: Optical biosensing methods (biolayer interferometry and plasmon surface resonance) were used to screen for interactions between HypA, HypB, UreE and UreG. Results: Using both methods, affinity constants were found to be 5 nM and 13 nM for HypA-UreE and 8 mu M and 14 mu M for UreG-UreE. Neither Zn2+ nor Ni2+ had an effect on the kinetics or stability of the HypA-UreE complex. By contrast, addition of Zn2+, but not Ni2+, altered the kinetics and greatly increased the stability of the UreE-UreG complex, likely due in part to Zn2+-mediated oligomerization of UreE. Finally our results unambiguously show that HypA, UreE and UreG cannot form a heterotrimeric protein complex in vitro; instead. HypA and UreG compete with each other for UreE recognition. eneral significance: Factors influencing the pathogen's nickel budget are important to understand pathogenesis and for future drug design. (c) 2012 Elsevier B.V. All rights reserved.