Temperature (T) dependence of kinetic isotope effects (KIEs) for H-tunneling reactions has been recently used to discuss about the role of protein dynamics in enzyme catalysis. Frequently observed weak T-dependence of KIEs in wild-type enzymes has been used to suggest narrowly distributed short donor-acceptor distances (DADs) caused by strong protein compressive vibrational effects in the H-tunneling coordinate. This strategy of study has been debated as simulations following other H-tunneling models sometimes gave alternative explanations. We recently reported the correlation of the donor-acceptor distances (DAD) distributions in the productive reactant complexes (PRC’s) and tunneling-ready states (TRS’s) to the T-dependence of KIEs of the NADH/NAD⁺ model reactions in various solvents. We found that a weaker T-dependence of KIEs happens in a more polar aprotic solvent in which the system has a shorter average DAD_PRC and DAD_TRS. In that paper, the DAD_PRC distributions were abstracted from the optimized stable PRC structures from the QM computations, and the DAD_TRS information was qualitatively evaluated from the kinetic rates including the secondary (2°) KIEs. Herein, we report the dynamical distribution of the fluctuating DAD_PRC’s in acetonitrile vs. chloroform (high vs. low polarity), which were obtained by MD computations. We also computed the TRS structures and fitted the hence computed 2° KIEs to the observed ones to derive the quantitative DAD_TRS information for the reactions in the two solvents. We found that the fluctuating DAD_PRC’s are more narrowly distributed and the average DAD_TRS is shorter in a more polar solvent of acetonitrile. Combining the observed weaker T-dependence of KIEs in acetonitrile, we conclude that a more strongly vibrational system corresponds with a weaker T-dependence of KIEs. Our results appear to support the explanation that links the weak T-dependence of KIEs to the donor-acceptor rigidity caused by strong enzyme compressive effects.

Mingxuan Bai

Appearances