Dihydrofolate reductase catalytic mechanism

Reaction Free Energy Profiles Using Free Energy Perturbation and Coordinate Coupling Methodologies Analysis of the Dihydrofolate Reductase Catalytic Mechanism... 

S. R. Stone and J. R. Morrison, Catalytic mechanism of the dihydrofolate reductase as... 

Because of its significance in cancer therapy and its small size, dihydrofolate reductase is one of the most studied of all enzymes. Numerous NMR studies385 387 and investigations of catalytic mechanism and of other properties388 have been conducted. Many mutant forms have been created 389-391 For example, substitution of Asp 27 (Asp 26 in L. casei) of the E. coli... 

Osborne, M., Schnell, J., Benkovic, S., Dyson, H. and Wright, P. (2001). Backbone dynamics in dihydrofolate reductase complexes role of loop flexibility in the catalytic mechanism. Biochemistry 40, 9846-9859... 

Geeady, J. E. (1985) Theoretical Studies on the Activation of the Pterin Cofactor in the Catalytic Mechanism of Dihydrofolate Reductase, Biochemistry 24, 4761-4766. 

Dihydrofolate reductase (DHFR, EC 1.5.1.3) is an essential enzyme required for normal folate metabolism in prokaryotes and eukaryotes. Its role is to maintain necessary levels of tetrahydrofolate to support the biosynthesis of purines, pyrimidines and amino acids. Many compounds of pharmacological value, notably methotrexate and trimethoprim, vork by inhibition of DHFR. Their clinical importance justified the study of DHFR in the rapidly evolving field of enzymology. Today, there is a vast amount of published literature (ca. 1000 original research articles) on the broad subject of dihydrofolate reductase contributed by scientists from diverse disciplines. We have selected kinetic, structural, and computational studies that have advanced our understanding of the DHFR catalytic mechanism with special emphasis on the role of the enzyme-substrate complexes and protein motion in the catalytic efficiency achieved by this enzyme. 

Castillo, R., Andres, J., Moliner, V. (1999) Catalytic mechanism of dihydrofolate reductase enzyme ... 

Cummins, P. L., Gready, J. E. (2001) Energetically most likely substrate and active-site protonation sites and pathways in the catalytic mechanism of dihydrofolate reductase, J. Am. Chem. Soc. 123, 3418-3428. 

Wagner, C. R., Thillet, J., Benkovig, S. J. (1992) Complementary Pertuba-tion of the Kinetic Mechanism and Catalytic Effectiveness of Dihydrofolate Reductase by Side-Chain Interchange, Biochemistry 31, 7834-7840. 

The effect of the electrostatic interactions on the catalytic mechanism of dihydrofolate reductase (DHFR) from E. coli has been undertaken by Cannon et al.130 The study of two enzyme-cofactor complexes and two enzyme-cofactor-substrate complexes shows that Asp-27 plays a major role in catalysis, and that the formation of 4-hydroxypterin is the more likely intermediate than... 

Ferrer S, Silla E, Tunon I, Marti S, Moliner V (2003) Catalytic mechanism of dihydrofolate reductase enz5nne. A combined quantum-mechanicaFmolecular-mechanical characterization of the N5 protonation step. J Phys Chem B 107(50) 14036-14041... 

Isotopes can be used in another way to measure the energy barrier heights for various steps in the catalytic mechanism as noted above for the reaction catalyzed by dihydrofolate reductase. For example, if a proton transfer is involved in the rate-limiting step, then substitution of that proton with one of the heavier isotopes of hydrogen (deuterium or tritium) will cause the step to proceed more slowly. These so-called kinetic isotope effect experiments in combination with steady-state rate measurements in the case of TIM allowed the elucidation of the rate constants for partitioning of the cw-enediol intermediate and construction of a detailed kinetic scheme as shown above for dihydrofolate reductase. 

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