DFT modeling of structures and redox potentials of wild-type, Nickel-substituted and mutated (N47S/M121L, HPAz) Azurin

Azurin (Az) from Pseudomonas aeruginosa is a redox active protein belonging to the family of cupredoxins. Cupredoxins span a wide range of reduction potentials (E0) going from stellacyanin having the lowest potential of ca. 184mV to rusticyanin showing the higher potential of ca. 680mV. Several works have been devoted to the understanding of the factors influencing E0 by changing primary coordination sphere ligands or exploring secondary coordination sphere mutations. To this goal, a series of Az mutants have been designed and showed that E0 could be tuned over a very broad range (between 90mV and 640mV) without significantly perturbing the metal binding site. Among these mutants, the HPAz variant showed the highest E0 value (970mV) ever reported for Az while a significant lowering of E° value (-590mV) has been reached in a Ni-substituted Az. In this paper, we computed the B3LYP energies and Gibbs free energies of oxidized and reduced models of wild-type Az, Ni-substituted Az and two mutants (N47S/M121L and HPAz) of the protein Az to estimate their E0. The results show that the employed strategy is able to reproduce the experimental lowering or increasing of E0 among the studied Az proteins