Crystallographic study of mutant Lys120Leu Xenopus laevis Cu,Zn superoxide dismutase

Abstract

Theoretical calculations and experimental measurements on the Xenopus laevis Cu,Zn superoxide dismutase (XSODB) wild-type protein and on some of its engineered mutants showed that the electrostatic arrangement around the active site channel plays a fundamental role in determining the catalytic properties of the enzyme. Lys120, which lies on the lip of the active site channel, about 11 Angstrom from the catalytic copper ion, influences the enzyme electrostatic environment and binding selectivity. Neutralization of this residue has the effect of decreasing the activity of the enzyme versus the negatively charged substrate. In order to get precise information about the mutated residue and its effects on the structure of the engineered protein, the crystal structure of single site Lys120Leu mutant XSODB was determined at 2.0 Angstrom resolution, and refined to an R-factor value of 0.181. The structure of Lys120Leu mutant XSODB is little affected by the amino-acid substitution, suggesting that the main effect-of the mutation is perturbation of the electrostatic properties of the SOD catalytic center