Pyridine Nucleotide Chemistry. A New Mechanism for the Hydroxide-Catalyzed Hydrolysis of the Nicotinamide-Glycosyl Bond

Article abstract of DOI:10.1021/ja00215a041

The mechanism of hydroxide-catalyzed hydrolysis of the glycosyl bond of β-NAD⁺ has been reinvestigated.The pH dependence of the rate of hydrolysis of β-NAD⁺ and related compounds has been determined over a range from pH 4.3 to 13.5.Between pH 8.5 and 11 the log of the rate constant is linearly dependent on pH with a slope of unity.Below pH 6.5 and above pH 12.5 the reaction becomes pH independent.The product of the reaction over the entire pH range is nicotinamide.A nonlinear least-squares fit of the data yields a pK_eq corresponding to the pK_a of ionization of the nicotinamide ribose diol, which has been determined independently by 13C NMR.Methanolysis of β-NAD⁺ yields a ratio of 3.7:1 for the β and α anomers of 1'-O-methyl-ADP ribose.Hydrolysis in a methanol/water mixture shows no selectivity for attack on the basis of the nucleophilicity of the attacking species.The importance of the ribose diol in the hydrolysis reaction was investigated with the isopropylidene derivative of β-nicotinamide riboside.Hydroxide-catalyzed decomposition of β-2',3'-O-isopropylidene nicotinamide riboside is pH independent below pH 7 and linearly dependent on hydroxide concentration above pH 10.In contrast to the results for β-NAD⁺, no pH-independent region is observed at high pH and the product of the pH-dependent reaction is 2-hydroxy-3-pyridinecarboxaldehyde; i.e., no detectable hydroxide-catalyzed release of nicotinamide is observed.On the basis of these data, as well as solvent isotope effects and data from previous investigations, we propose a new mechanism in which dissociative cleavage of nicotinamide-glycosyl bond is facilitated by the nicotinamide ribose diol anion through noncovalent stabilization of an oxo carbocation intermediate.