Computational Mutation Analysis of Enzymatic Reaction

Abstract

Density functional theory (DFT) calculations are established as a useful research tool to investigate the structures and reactivity of biological systems; however, their high computational costs still restrict their applicability to systems of several tens up to a few hundred atoms. Recently, a combined quantum mechanical/molecular mechanical (QM/MM) approach has become an important method to study enzymatic reactions. In the past several years, we have investigated B12-dependent diol dehydratase using QM/MM calculations. The enzyme catalyzes chemically difficult reactions by utilizing the high reactivity of free radicals. In this paper, we explain our QM/MM calculations for the structure and reactivity of diol dehydratase and report key findings with respect to the catalytic roles of the active-site amino acid residues, computational mutational analysis of the active-site amino acid residues, assignment of the central metal ion, and function of the central metal ion. Our QM/MM calculations can correctly describe the structures and activation barriers of intermediate and transition states in the protein environment. Moreover, predicted relative activities of mutants are consistent with experimentally observed reactivity. These results will encourage the application of QM/MM research to the mechanistic study of enzymatic reactions, functional analysis of active-site residues, and rational design of enzymes with new catalytic functions.