Unveiling the Catalytic Mechanism of NADP+-Dependent Isocitrate Dehydrogenase with QM/MM Calculations

Abstract

We have determined the catalytic mechanism of the human cytosolic homodimeric isocitrate dehydrogenase (hICDH), an enzyme involved in the regulation of tumorogenesis. Our study constitutes the first theoretical attempt to describe the entire catalytic cycle of hICDH. In agreement with earlier experimental proposals, the catalysis was shown to proceed in three steps: (1) NADP+ reduction by the isocitrate substrate with the help of the Lys212B base, (2) β-decarboxylation of the resulting oxalosuccinate, generating an enolate, and (3) protonation of this intermediate by Tyr139A, giving rise to the α-ketoglutarate product. Our study supports that the β-decarboxylation of oxalosuccinate is the most likely rate-limiting step, with an activation Gibbs free energy of 16.5 kcal mol–1. The calculated values are in close agreement with the 16–17 kcal mol–1 range, derived by the application of transition state theory to the reaction rates determined experimentally (11 to 38 s–1). We emphasize the role of Mg2+ and Asp...