The Journal of Biological Physics and Chemistry

Dynamic and stochastic patterns of enzyme action. A key to the interpretationof "normal" and "small" solvent kinetic isotope effects (k^H/k^D)

Dimitri E. Khoshtariya, Mikhael Shushanyan, Rusudan Sujashvili and Tina D. Dolidze

Available experimental data on the solvent kinetic isotope and viscosity effects for comparable enzyme-catalysed hydrolytic processes are discussed and rationalized on the basis of fundamental equations of modern charge-transfer theory. It is shown that the manifestation of the primary kinetic isotope effect (KIE) connected with proton transfer in the course of the rate-determining catalytic step points to an intrinsic nonadiabatic mechanism excluding the possibility of detection of Kramers-type viscosity dependence. In turn, observation of Kramers-type viscosity dependence of the catalytic constant strongly suggests an adiabatic rather than a nonadiabatic physical mechanism, allowing only for the solvent kinetic isotope effect due to the different viscosities of the applied solvents (H₂O and D₂O). Hence, the key prediction of the modern charge-transfer theory, mutual exclusion of the primary kinetic isotope effect and Kramers-type viscosity dependence is shown to be justified. The conclusions of the present analysis are in accord with the results of recent studies on the intrinsic charge-transfer mechanism changeover from nonadiabatic to adiabatic for the case of a redox-active protein, cytochrome c, operating at Au/SAM junctions.

Keywords: adiabatic/nonadiabatic mechanisms, carbonyl group, charge-transfer theory, enzymatic hydrolysis, general base catalysis, kinetic isotope effects, proton tunnelling, rate constants, tetrahedral intermediate, viscosity effects