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.