On interpreting the inhibition of and catalysis by dihydrofolate reductase

Inhibition of dihydrofolate reductase by methotrexate requires the presence of the p-aminobenzoyl glutamate side chain to achieve an isomerization of the initial encounter complex to the final tightly bound enzyme species. Site-specific mutagenesis at Phe31 revealed that hydrophobic interactions at this residue alone may contribute ∼ 2.5 kcal mol^-1 to the binding energy of the drug. This binding perturbation extends to the folate substrates but with a minimal (less than twofold) effect on the chemical step in turnover owing to a uniform effect on ground and transition states.