Molecular Interactions of Competitive Inhibitors with Bovine Liver Glutamate Dehydrogenase

Four structural analogues of l-glutamate (glutaric acid, thiodiglycolic acid, oxydiglycolic acid, and iminodiacetic acid) were tested for inhibition as a function of pH, of coenzyme NADP+ or NAD+, and of their electronic properties as determined from molecular orbital theory considerations. Plots of pVmax and p(Km/Vmax) versus pH for the uninhibited and inhibited reaction of the oxidative deamination of l-glutamate with NADP+ indicated an ionizable group or groups in the enzyme complex with a pK of 7.7 to 7.8. Double reciprocal plots of velocity versus glutamate concentrations in the presence of NADP+ or NAD+ showed that each structural analogue of glutamate was a competitive inhibitor with respect to glutamate. NAD+ increased the affinity of inhibitor for enzyme in comparison with NADP+. The relative effectiveness of each inhibitor as determined by apparent Ki values, calculated from Dixon plots, was correlated inversely with the absolute sigma charge density of the meso atom of the inhibitor. This suggested that desolvation of this atom may be important for the combination of inhibitor with enzyme. A high degree of solvation as indicated by the magnitude of absolute charge density could have decreased the interaction of inhibitor with enzyme.