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Loveridge, EJ, Tey, LH and Allemann, RK (2010) Solvent effects on catalysis by Escherichia coli dihydrofolate reductase. J. Am. Chem. Soc. 132:1137-43


Hydride transfer catalyzed by dihydrofolate reductase (DHFR) has been described previously within an environmentally coupled model of hydrogen tunneling, where protein motions control binding of substrate and cofactor to generate a tunneling ready conformation and modulate the width of the activation barrier and hence the reaction rate. Changes to the composition of the reaction medium are known to perturb protein motions. We have measured kinetic parameters of the reaction catalyzed by DHFR from Escherichia coli in the presence of various cosolvents and cosolutes and show that the dielectric constant, but not the viscosity, of the reaction medium affects the rate of reaction. Neither the primary kinetic isotope effect on the reaction nor its temperature dependence were affected by changes to the bulk solvent properties. These results are in agreement with our previous report on the effect of solvent composition on catalysis by DHFR from the hyperthermophile Thermotoga maritima. However, the effect of solvent on the temperature dependence of the kinetic isotope effect on hydride transfer catalyzed by E. coli DHFR is difficult to explain within a model, in which long-range motions couple to the chemical step of the reaction, but may indicate the existence of a short-range promoting vibration or the presence of multiple nearly isoenergetic conformational substates of enzymes with similar but distinct catalytic properties.


PubMed Online version:10.1021/ja909353c


Catalysis; Escherichia coli/enzymology; Hydrogen-Ion Concentration; Kinetics; Models, Molecular; Protein Conformation; Solvents/chemistry; Temperature; Tetrahydrofolate Dehydrogenase/chemistry; Tetrahydrofolate Dehydrogenase/metabolism; Thermotoga maritima/enzymology


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