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 Ni(2+) 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 5nM and 13nM for HypA-UreE and 8μM and 14μM for UreG-UreE. Neither Zn(2+) nor Ni(2+) had an effect on the kinetics or stability of the HypA-UreE complex. By contrast, addition of Zn(2+), but not Ni(2+), altered the kinetics and greatly increased the stability of the UreE-UreG complex, likely due in part to Zn(2+)-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.


CONCLUSIONS
Factors influencing the pathogen's nickel budget are important to understand pathogenesis and for future drug design.
Department of Microbiology, University of Georgia, 805 Biological Sciences Bldg., Athens, GA, USA.
Stéphane L
SL
Benoit
Jonathan L
JL
McMurry
Stephanie A
SA
Hill
Robert J
RJ
Maier
D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive / Q000502:physiology*; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D016480:Helicobacter pylori / Q000201:enzymology / Q000378:metabolism*; D006864:Hydrogenase / Q000378:metabolism; D056511:Metallochaperones; D028044:Phosphate-Binding Proteins; D011485:Protein Binding; D055503:Protein Multimerization / Q000502:physiology; D013057:Spectrum Analysis / Q000379:methods; D020349:Surface Plasmon Resonance / Q000379:methods; D014510:Urease / Q000378:metabolism
D001426:Bacterial Proteins; D002352:Carrier Proteins; C578705:HypA protein, Helicobacter pylori; C089637:HypB protein, bacteria; D056511:Metallochaperones; D028044:Phosphate-Binding Proteins; C090404:UreE protein, Bacteria; C082270:ureG protein, Bacteria; D006864:Hydrogenase; D014510:Urease; D019204:GTP-Binding Proteins