Rationalization of the Enantioselectivity of Subtilisin in DMF

Abstract

Herein we examine the origin of enantioselectivity in the serine protease subtilisin in DMF through the use of molecular dynamics (MD) and free energy perturbation (FEP) simulations. In particular, we are interested in the resolution of a racemic mixture of sec-phenethyl alcohol by a transesterification reaction with the acylating agent vinyl acetate, catalyzed by subtilisin in anhydrous dimethylformamide (DMF). To study the enantioselectivity in this case, we examined the tetrahedral intermediate as a model of the enzyme transition state (as has been done in the past). A critical aspect of this study was the determination of the charge distribution of the two (R and S) tetrahedral intermediates through the use of a combined quantum mechanical/molecular mechanical electrostatic potential fitting methodology. In designing the active site charge model, we found that the R and S tetrahedral intermediates have significantly different charge distributions due to the presence of the stereodifferentiating enviro...