Catalytic Roles of Histidine and Arginine in Pyruvate Class II Aldolase: A Perspective from QM/MM Metadynamics

The retro-aldol reaction catalyzed by pyruvate class II aldolase is investigated with QM/MM metadynamics; this enzyme transforms the substrate of 4-hydroxy-2-ketoacid into pyruvate and aldehyde through the aldol cleavage. The hydroxyl group of the substrate is deprotonated by His45 with the aid of the metal-bound water, while the metal-bound hydroxide proposed in the literature is observed as a transient species. The deprotonation appears to enhance substrate binding between the deprotonated substrate and the active site. The reactive alkoxide is further stabilized by the salt bridge of Arg70-Asp42, facilitating the following aldol cleavage. The simulations show that the C−C bond cleavage is the rate-determining step, and the calculated barrier of approximately 14 kcal mol-1 agrees reasonably with experimental data.