10.1021/acscatal.9b03345.s001 Philip Pagano Philip Pagano Qi Guo Qi Guo Chethya Ranasinghe Chethya Ranasinghe Evan Schroeder Evan Schroeder Kevin Robben Kevin Robben Florian Häse Florian Häse Hepeng Ye Hepeng Ye Kyle Wickersham Kyle Wickersham Alán Aspuru-Guzik Alán Aspuru-Guzik Dan T. Major Dan T. Major Lokesh Gakhar Lokesh Gakhar Amnon Kohen Amnon Kohen Christopher M. Cheatum Christopher M. Cheatum Oscillatory Active-Site Motions Correlate with Kinetic Isotope Effects in Formate Dehydrogenase American Chemical Society 2019 FDH exhibits oscillatory frequency fluctuations temperature dependence transition state structure Kinetic Isotope Effects Formate IR mutations influence active-site dynamics enzyme-catalyzed hydride transfers Oscillatory Active-Site Motions Correlate hydride transfer reaction Candida boidinii exhibits KIE 2019-11-11 05:29:43 Journal contribution https://acs.figshare.com/articles/journal_contribution/Oscillatory_Active-Site_Motions_Correlate_with_Kinetic_Isotope_Effects_in_Formate_Dehydrogenase/10282325 Thermal motions of enzymes have been invoked to explain the temperature dependence of kinetic isotope effects (KIEs) in enzyme-catalyzed hydride transfers. Formate dehydrogenase (FDH) from Candida boidinii exhibits a temperature-independent KIE that becomes temperature-dependent upon mutation of hydrophobic residues in the active site. Ternary complexes of FDH that mimic the transition state structure allow investigation of how these mutations influence active-site dynamics. A combination of X-ray crystallography, two-dimensional infrared (2D IR) spectroscopy, and molecular dynamic simulations characterize the structure and dynamics of the active site. FDH exhibits oscillatory frequency fluctuations on the picosecond timescale, and the amplitude of these fluctuations correlates with the temperature dependence of the KIE. Both the kinetic and dynamic phenomena can be reproduced computationally. These results provide experimental evidence for a connection between the temperature dependence of KIEs and motions of the active site in an enzyme-catalyzed reaction consistent with activated tunneling models of the hydride transfer reaction.