Electrochemical, Spectroscopic, and Density Functional Theory Characterization of Redox Activity in Nickel-Substituted Azurin: A Model for Acetyl-CoA Synthase.

Nickel-containing enzymes are key players in global hydrogen, carbon dioxide, and methane cycles. Many of these enzymes rely on NiI oxidation states in critical catalytic intermediates. However, due to the highly reactive nature of these species, their isolation within metalloenzymes has often proved elusive. In this report, we describe and characterize a model biological Ni1 species that has been generated within the electron transfer protein, azurin. Replacement o f the native copper cofactor with nickel is shown to preserve the redox activity o f the protein. The NiII/I couple is observed at -590 mV versus NHE, with an interfacial electron transfer rate of 70 s-1. Chemical reduction of NiIIAz generates a stable species with strong absorption features at 350 nm and a highly anisotropic, axial EPR signal with principal g-values of 2.56 and 2.10. Density functional theory calculations provide insight into the electronic and geometric structure o f the NiI species, suggesting a trigonal planar coordination environment 'The predicted spectroscopic features o f this low-coordinate nickel site are in good agreement with the experimental data. Molecular orbital analysis suggests potential for both metal-centered and ligand-centered reactivity, highlighting the covalency of the metal--thiolate bond. Characterization of a stable NiI species within a model protein has implications for understanding the mechanisms o f complex enzymes, including acetyl coenzyme A synthase, and developing scaffolds for unique reactivity. [ABSTRACT FROM AUTHOR]