The Catalytic Power of Dihydrofolate Reductase and Other Enzymes Arises from Electrostatic Preorganization, Not Conformational Motions

This is an accepted version of a paper published in Proceedings of the National Academy of Sciences of the United States of America. This paper has been peer-reviewed but does not include the final publisher proof-corrections or journal pagination. "Catalysis by dihydrofolate reductase and other enzymes arises from electrostatic preorganization, not conformational motions" Access to the published version may require subscription. 2 ABSTRACT The proposal that enzymatic catalysis is due to conformational fluctuations has been previously promoted by means of indirect considerations. However, recent works have focused on cases where the relevant motion can be described in terms of rather unique conformational states, whose population could be manipulated by mutations. In particular, a recent work has claimed to provide direct experimental evidence for a dynamical contribution to catalysis in dihydrofolate reductase, where blocking a relevant conformational coordinate was identified as suppressing the motion toward the occluded conformation. The present work utilizes computer simulations to elucidate the true molecular basis for the experimentally observed effect. We start by reproducing the trend in the measured change in the catalytic effect of the enzyme upon mutations (that were assumed to create a " dynamical knockout "), by calculating the change in the corresponding activation barriers, and without the need to invoke dynamical effects. We then generate the catalytic landscape of the enzyme and demonstrate that motions in the conformational space do not help drive catalysis. We also discuss the role of flexibility and conformational dynamics in catalysis, once again demonstrating that their role is negligible and that the largest contribution to catalysis arises from electrostatic preorganization. Finally, we point out that the changes in the reaction potential surface change the reorganization free energy (which includes entropic effects), and such changes in the surface also alter the motion.