Multiple Reaction Pathways in the Morphinone Reductase-Catalyzed Hydride Transfer Reaction

[Image: see text] Morphinone reductase (MR) is an important model system for studying the contribution of protein motions to H-transfer reactions. In this research, we used quantum mechanical/molecular mechanics (QM/MM) simulation together with transition path sampling (TPS) simulation to study two important topics of current research on MR: the existence of multiple catalytic reaction pathways and the involvement of fast protein motions in the catalytic process. We have discovered two reaction pathways for the wild type and three reaction pathways for the N189A mutant. With the committor distribution analysis method, we found reaction coordinates for all five reaction pathways. Only one wild-type reaction pathway has a rate-promoting vibration from His186, while all of the other four pathways do not involve any protein motions in their catalytic process through the transition state. The rate-promoting vibration in the wild-type MR, which comes from a direction perpendicular to the donor–acceptor axis, functions to decrease the donor–acceptor distance by causing a subtle “out-of-plane” motion of a donor atom. By comparing reaction pathways between the two enzymes, we concluded that the major effect of the N189A point mutation is to increase the active site volume by altering the active site backbone and eliminating the Asn189 side chain. This effect causes a different NADH geometry at the reactant state, which very well explains the different reaction mechanisms between the two enzymes, as well as the disappearance of the His186 rate-promoting vibrations in the N189A mutant. The unfavorable geometry of the NADH pyridine ring induced by the N189A point mutation is the potential cause of multiple reaction pathways in N189A mutants.