Mechanistic study of the biosynthesis of R-phenylcarbinol by acetohydroxyacid synthase enzyme using hybrid quantum mechanics/molecular mechanics simulations.

The biosynthesis of R-phenylacetylcarbinol (R-PAC) by the acetohydroxy acid synthase, (AHAS) is addressed by molecular dynamics simulations (MD), hybrid quantum mechanics/molecular mechanics (QM/MM), and QM/MM free energy calculations. The results show the reaction starts with the nucleophilic attack of the C2α atom of the HEThDP intermediate on the C atom of the carbonyl group of benzaldehyde substrate via the formation of a transition state (TS1) with the HEThDP intermediate under 4'-aminopyrimidium (APH) form. The calculated activation free energy for this step is 17.4kcal mol at 27 °C. From this point, the reaction continues with the abstraction of H atom of the HEThDP intermediate by the O atom of benzaldehyde to form the intermediate I. The reaction is completed with the cleavage of the bond C2α-C2 to form the product R-PAC and to regenerate the ylide intermediate under the APH form, allowing in this way to reinitiate to the catalytic cycle once more. The calculated activation barrier for this last step is 15.9kcal mol at 27 °C.