Evidence for binding of NAD dimers to NAD-dependent dehydrogenases.

The binding of dimers of NAD, (NAD)2, to lactate, malate and alc. dehydrogenases was studied by the fluorescence quenching technique. Whereas alc. dehydrogenase shows a low binding ability, malate and lactate dehydrogenases bind (NAD)2 in a specific way with high affinity. Malate dehydrogenase binds (NAD)2 more than NADH. All 3 dehydrogenases are inhibited by (NAD)2, which behaves as a competitive inhibitor with respect to both NAD and NADH. These results show that (NAD)2 is bound to the nucleotide-specific binding site of the dehydrogenases. (NAD)2 stoichiometrically reacts with ferricyanide at variance with NADH. The specific interactions with the NAD-dependent dehydrogenases and the ability to enter in monoelectronic redox cycles suggest possible physiol. roles for (NAD)2.