Anomalous Equilibrium and Kinetic α-Deuterium Secondary Isotope Effects Accompanying Hydride Transfer from Reduced Nicotinamide Adenine Dinucleotide

Article abstract of DOI:10.1021/ja00532a035

The kinetic α-deuterium secondary isotope effect on the second-order rate constant has been measured for the nonenzymatic direct hydride transfer reduction of 4-cyano-2,6-dinitrobenzenesulfonate by NADH (deuterium substitution of the hydrogen bonded to the 4 carbon of NADH which is not transferred to the acceptor).Values of 1.156 +/- 0.018 and 1.1454 +/- 0.0093 were obtained using direct and intramolecular competition methods, respectively.The corresponding (enzyme catalyzed) equilibrium isotope effects were found to be 1.013 +/- 0.020 and 1.0347 +/- 0.0087 as determined by direct and intermolecular competition methods, respectively.Thus, the value of the kinetic effect is significantly greater than that on the equilibrium.It is suggested that this may arise either from participation of the α hydrogen in a hyperconjugative stabilization of an early transition state or from its participation in the reaction coordinate motion of a nonlinear activated complex.The values of the equilibrium effect allow calculation of a fractional factor (relative to acetylene) for hydrogen bonded to the 4 carbon of NAD⁺ of 1.448 +/- 0.028 or 1.418 +/- 0.020.This is larger than expected based on comparison with hydrogen bound of sp² carbon in propene (1.136) or benzene (1.368) but is consistent with the decreased aromatic character of pyridinium vibrational spectra.The lack of a significant inverse value for the equilibrium α-deuterium effect suggests complications in the interpretation of reported kinetic secondary effects of 0.85 and 1.2 for the forward (sp³ -> sp%&2) and reverse (sp² -> sp³) rate constants for the nonenzymatic transhydrogenation of N-benzyl-1,4-dihydronicotinamide and its nicotinamide salt.