The use of 3-acetyl-pyridine adenine dinucleotide in the spectrophotometric assay of enzymatic activities coupled to dehydrogenases of unfavorable equilibrium

The high redox potential of the couple $\text{NAD}{}^{\mathrm{+}}\text{NADH}$ has limited in the past the application of dehydrogenases in the coupled assay of enzymes. Here the way to obviate this difficulty is presented, by using the chemical analog 3-acetyl-NAD⁺ and an auxiliary enzyme after the dehydrogenase. A general kinetic scheme of the multienzyme systems is presented in the discussion that helps to explain why this approach succeeds in making these dehydrogenases satisfactory auxiliary enzymes even in the unfavorable direction.