Nickel binding shifts Helicobacter pylori HypA toward compact conformations

Helicobacter pylori ( Hp ) is a Gram-negative human pathogen that relies on the nickel enzymes urease and Ni,Fe‑hydrogenase for gastric colonization. Delivery of Ni(II) to the active sites of these enzymes is mediated by the metallochaperone Hp HypA, which contains a high-affinity structural Zn(II) site and a lower-affinity Ni(II) site, in two distinct domains. Although the NMR solution structure of the apo form Zn- Hp HypA is known, the impact of Ni(II) binding on the structure remains unclear. Here, NMR 15 N relaxation experiments have been applied to probe the backbone internal dynamics and the rotational diffusion of Zn- Hp HypA and Ni,Zn- Hp HypA. Residue-resolved model-free analysis revealed broadly similar ps–ns backbone motional amplitudes in the apo and holo states. In contrast, Ni(II) binding increased rotational diffusion, indicating a reduced hydrodynamic radius and a shift toward a more compact conformational ensemble. These observations support a model in which Ni(II) binding generates a delivery-competent protein structure by narrowing the accessible conformational distribution during partner recognition. Previously, calorimetric data reported on the interaction between Hp HypA and the urease metallochaperone homodimeric Hp UreE 2 , which showed micromolar Ni(II) binding to Hp HypA but the emergence of a sub-nanomolar, strongly exothermic Ni site in the HypA•UreE 2 complex. The observed nickel-induced ensemble compaction determined by solution NMR spectroscopy provides a plausible physical basis for Ni-dependent conformational switching to promote high-affinity protein-protein recognition. Interpretation of the compaction of Helicobacter pylori HypA upon binding Ni(II). • Ni(II) binding compacts Helicobacter pylori HypA without altering internal backbone dynamics. • Ni(II) binding increases the rotational diffusion of Hp HypA consistent with a shift toward a more compact conformational ensemble. • The structural compaction is consistent with tighter packing of a helical structural motif upon Ni(II) binding. • The nickel-induced compact state likely prepares Hp HypA for nickel delivery and partner recognition, providing a physical basis for the very high-affinity Hp HypA• Hp UreE 2 interaction in the Ni-bound system.