INTERACTION AND ACTIVITIES OF NAD+ AND NAD+ ANALOGS WITH FOUR DEHYDROGENASES

Abstract

The effects of 6 NAD analogs on the Km, Vmax, KD, fluorescence quenching, k1, and decompn. of the enzyme-NADH complex with 4 dehydrogenases are reported. The interaction of the adenine ribose and adenine of NAD of reactions done in soln., where catalysis is complete is compared with the x-ray maps where catalysis is not complete. It appears that the 2'- and 3'-hydroxyl groups of NAD are not essential for bonding, but are essential for proper orientation of the nicotinamide ring to produce a productive complex. NAD, 2'-deoxy-NAD (2'-dNAD) and 3'-deoxy-NAD (3'-dNAD) have the same affinity with the enzymes. Possibly the 2'-hydroxyl group of NAD does not result in a typical 3-5 kcal/mol hydroxyl-carboxylate H-bond. Alternatively, 2'- and 3'-dNAD may not be positioned correctly in the coenzyme domain for max. catalysis. This requires that the intrinsic binding energy for NAD, 2'- and 3'-dNAD be the same. Because yeast alc. dehydrogenase and presumably horse liver alc. dehydrogenase, glyceraldehyde phosphate dehydrogenase, and lactate dehydrogenase have compulsory ordered kinetics, a decrease in both k1 and the rate of decompn. of the enzyme-NADH complex was obsd. Replacement of adenosine of NAD with tubercidin, formycin, inosine, or etheno-adenosine produced changes in the Km, KD, and Vmax that are discussed in terms of their accommodations within the coenzyme domain. [on SciFinder(R)]