Solvent Effects on Catalysis by Escherichia coli Dihydrofolate Reductase

摘要

Abstract: Hydride transfer catalyzed by dihydrofolate reductase (DHFR) has been described previouslynwithin an environmentally coupled model of hydrogen tunneling, where protein motions control bindingnof substrate and cofactor to generate a tunneling ready conformation and modulate the width of thenactivation barrier and hence the reaction rate. Changes to the composition of the reaction medium arenknown to perturb protein motions. We have measured kinetic parameters of the reaction catalyzed bynDHFR from Escherichia coli in the presence of various cosolvents and cosolutes and show that thendielectric constant, but not the viscosity, of the reaction medium affects the rate of reaction. Neithernthe primary kinetic isotope effect on the reaction nor its temperature dependence were affected bynchanges to the bulk solvent properties. These results are in agreement with our previous report on theneffect of solvent composition on catalysis by DHFR from the hyperthermophile Thermotoga maritima.nHowever, the effect of solvent on the temperature dependence of the kinetic isotope effect on hydridentransfer catalyzed by E. coli DHFR is difficult to explain within a model, in which long-range motionsncouple to the chemical step of the reaction, but may indicate the existence of a short-range promotingnvibration or the presence of multiple nearly isoenergetic conformational substates of enzymes withnsimilar but distinct catalytic properties.