Journal:JBIC:23

Selectivity of Ni(II) and Zn(II) binding to Sporosarcina pasteurii UreE, a metallo-chaperone in the urease assembly: a calorimetric and crystallographic study

Barbara Zambelli, Katarzyna Banaszak, Anna Merloni, Agnieszka Kiliszek, Wojciech Rypniewski, Stefano Luciano Ciurli ^[1]

Molecular Tour
Urease is a nickel-dependent enzyme that plays a critical role in the biogeochemical nitrogen cycle by catalyzing the hydrolysis of urea to ammonia and carbamate. This enzyme, initially synthesized in the apo-form, needs to be activated by nickel ion incorporation into the active site, driven by the dimeric metallo-chaperone UreE. The present study explores the metal selectivity and affinity of UreE from Sporosarcina pasteurii for cognate (Ni(II)) and non-cognate (Zn(II)) metal ions. The , polypeptide chain A and B are shown in green and darkmagenta respectively, Ni ion shown as a cyan ball, Zn ion shown as a grey ball, two His100 shown as magenta sticks. . In particular, the thermodynamic parameters of SpUreE for Ni(II) and Zn(II) binding have been determined using isothermal titration calorimetry. These experiments show that two Ni(II) ions bind to the protein dimer with positive cooperativity, with a high affinity and a low affinity site. Zn(II) binding to the protein, occurring in the same region and with similar affinity, causes metal-driven dimerization of the protein dimer. The crystal structure of the protein obtained in the presence of equimolar amounts of both metal ions indicates that the high affinity metal binding site preferentially binds Ni(II) over Zn(II). The ability of the protein to select Ni(II) over Zn(II) was confirmed by competition experiments in solution as well as by analysis of X-ray anomalous dispersion data. Overall, the thermodynamics and structural parameters that modulate the metal ion specificity of different binding sites on the protein surface have been established.