Free Energy Landscape and Mechanistic Pathways of Transimination Reaction at the Active Site of Ornithine Decarboxylase Using Hybrid QM/MM Simulations

Transimination is a crucial biochemical process where a pyridoxal 5'-phosphate (PLP)-substrate complex is formed from a PLP-enzyme complex. This transformation is essential as it initiates the catalytic reaction of the enzyme, enabling the enzyme to perform its specific biochemical function. In this study, we have investigated the mechanistic details of transimination reaction at the active site of ornithine decarboxylase (ODC) using QM/MM simulations and enhanced sampling techniques. It is found that, in the first step, the backbone amino group nitrogen of the ornithine attacks the imine bond carbon of the PLP-enzyme complex, leading to the formation of a geminal diamine complex among PLP, lysine, and the substrate (ornithine). Our calculations reveal that the formation of this initial geminal diamine intermediate (GDI1) involves a free energy barrier of 4 kcal/mol. Following the formation of GDI1, a proton transfer takes place from the ornithine backbone nitrogen to the lysine69 side chain nitrogen, resulting in the formation of a second geminal diamine complex (GDI2). Although there are multiple possible pathways for this proton transfer to take place, our calculations show that the proton transfer at ODC occurs via the phenolic oxygen (rather than water) with appropriate changes in the dihedral angle facilitating this process. For the formation of the external aldimine from GDI2, we found a free energy barrier of 6 kcal/mol. The proton transfer step involves a free energy barrier of 10 kcal/mol, hence acts as the rate-limiting step in the transimination process.