A Kinetic isotope effect in purinic 8‐CH exchange of guanine nucleotides: Measurement by Raman spectroscopy

Abstract

AbstractRates of deuterium exchange of the 8‐CH groups of 5′‐rGMP and cGMP in D2O solutions (KH) are compared with rates of protium exchange of the 8‐CD groups of the same compounds in H2O solutions (KD). The ratio of the two rate constants (kH/kD) defines an isotope effect which is a combination of primary kinetic and solvent isotope effects, the former predominating. The net isotope effect was measured by Raman spectrophotometry over the temperature range 30–80°C. The Arrhenius activation energies (Ea) and pre‐exponential frequency factors (A) were also determined. The average values of kH/kD are 1.82 ± 0.15 for 5′‐rGMP and 1.43 ± 0.06 for cGMP. The relatively small kinetic isotope effects are indicative typically of an unsymmetrical transition state. The ratio E/E is unity in both compounds, indicating an early or reactant‐like transition state. The present data are consistent with, and extend the range of, the Bell–Goodall curve relating log (kH/kD) to ΔpK of the two acid–base systems involved in the proton transfer during the rate‐determining step.