Semiempirical calculations of the oxygen equilibrium isotope effect on binding of oxamate to lactate dehydrogenase.

Semiempirical methods have been used in an attempt to predict theoretically the experimentally observed value of 0.9840 for the oxygen isotope effect on binding of oxamate to lactate dehydrogenase. The overall strategy involved vibrational analysis of oxamate in two different environments; that of the active site residues and in aqueous solution. The comparison of calculated values with the experimentally determined isotope effect proved the AM1 Hamiltonian to be superior to the PM3 Hamiltonian in this modelling. While most tested methods of accounting for solvent effects on the vibrational frequencies of the solute yielded similar results it turned out that what was crucial for the purpose of determination of the isotope effect was the model of oxamate in the active site of the enzyme. In particular, the major factor responsible for the inverse value of this isotope effect can be ascribed to the formation of an ordered, bifurcated hydrogen bond between the oxamate carboxylate and the guanidinium group of the active site histidine.