cs9b03345_si_001.pdf (1.17 MB)
Oscillatory Active-Site Motions Correlate with Kinetic Isotope Effects in Formate Dehydrogenase
journal contribution
posted on 2019-11-11, 05:29 authored by Philip Pagano, Qi Guo, Chethya Ranasinghe, Evan Schroeder, Kevin Robben, Florian Häse, Hepeng Ye, Kyle Wickersham, Alán Aspuru-Guzik, Dan T. Major, Lokesh Gakhar, Amnon Kohen, Christopher M. CheatumThermal
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.