Zinc–Homocysteine binding in cobalamin‐dependent methionine synthase and its role in the substrate activation: DFT, ONIOM, and QM/MM molecular dynamics studies

Cobalamin‐dependent methionine synthase (MetH) is an important metalloenzyme responsible for the biosynthesis of methionine. It catalyzes methyl transfer from N5‐methyl‐tetrahydrofolate to homocysteine (Hcy) by using a zinc ion to activate the Hcy substrate. Density functional theory (B3LYP) calculations on the active‐site model in gas phase and in a polarized continuum model were performed to study the Zn coordination changes from the substrate‐unbound state to the substrate‐bound state. The protein effect on the Zn2+ coordination exchange was further investigated by ONIOM (B3LYP:AMBER)‐ME and EE calculations. The Zn2+‐coordination exchange is found to be highly unfavorable in the gas phase with a high barrier and endothermicity. In the water solution, the reaction becomes exothermic and the reaction barrier is drastically decreased to about 10.0 kcal/mol. A considerable protein effect on the coordination exchange was also found; the reaction is even more exothermic and occurs without barrier. The enzyme was suggested to constrain the zinc coordination sphere in the reactant state (Hcy‐unbound state) more than that in the product state (Hcy‐bound state), which promotes ligation of the Hcy substrate. Molecular dynamics simulations using molecular mechanics (MM) and PM3/MM potentials suggest a correlation between the flexibility of the Zn2+‐binding site and regulation of the enzyme function. Directed in silico mutations of selected residues in the active site were also performed. Our studies support a dissociative mechanism starting with the ZnO(Asn234) bond breaking followed by the ZnS(Hcy) bond formation; the proposed associative mechanism for the Zn2+‐coordination exchange is not supported. © 2011 Wiley Periodicals, Inc. J Comput Chem, 2011