Donor-Acceptor Separation Augments Temperature Dependence of Kinetic Isotope Effects in NADH Model Hydride Transfer...

Observed shift from temperature (T)-independent hydrogen tunneling kinetic isotope effects (KIEs) in enzymes to T-dependent KIEs in mutants has been attributed to the donor(D)-acceptor(A) separation effects caused by the weakened protein dynamical compression of D-A distances (DADs) in mutants. To examine the relationship between D-A separations (DADs) and T-dependence of KIEs (represented by ΔE_a = E_aD - E_aH), we design hydride-transfer reactions in solution. Our hypothesis is that a looser nucleus-transfer system exhibits a larger ΔE_a value. Herein, the ΔE_a's were determined for three series of apparent hydride-transfer reactions of NADH models in acetonitrile. These include hydride-transfers (1) from Hantzsch ester to 10-methyl-9-substituted(R)acridinium ions (RMA⁺), (2) from the reduced RMA⁺ (RMAH) to a benzoquinone structure, and (3) from RMAH to the [(BnTPEN)Fe(IV)═O]²⁺ complex. Reactions (2) and (3) use multistep electron-proton-electron sequential transfer mechanisms. ΔE_a increases from reactions (1) (0.94-1.19 kcal/mol) to (2) (1.14-1.60 kcal/mol) to (3) (3.05-5.05 kcal/mol), and within each series, ΔE_a increases with the size of the R substituent. The unusually high ΔE_a's observed for the iron(IV)-oxo complex reactions are likely partly attributed to electrostatic repulsion between like-charged RMAH^+• acid and [(BnTPEN)Fe(III)═O]⁺ in the reaction complex. These results support our hypothesis and the proposed role of protein dynamics in barrier compression for enzyme catalysis.