Understanding the reaction mechanism and intermediate stabilization in mammalian serine racemase using multiscale quantum-classical simulations.

Serine racemase (SerR) is a pyridoxal-5'-phosphate (PLP)-dependent enzyme catalyzing the racemization of l-Ser to d-Ser. In mammals, d-Ser is an endogenous coagonist required for the activation of N-methyl-d-aspartate receptors (NMDARs), thus making SerR a promising pharmaceutical target. However, mechanistic studies of SerR are scarce, and the details of the enzymatic racemization reaction are not fully understood. In the current study we elucidate the catalytic mechanism in SerR by employing combined multiscale classical/quantum simulations. The free energy profile of a model SerR racemization reaction is first calculated in the gas phase and in aqueous solution. To obtain the free energy profile for the enzymatic reaction, hybrid quantum mechanics/molecular mechanics molecular dynamics simulations in conjunction with umbrella sampling are performed. The results suggest that in SerR, similarly to the related enzyme alanine racemase, the unprotonated PLP-substrate intermediate is stabilized mostly due to solvation effects contributed by water molecules and active-site residues, as well as long-range electrostatic interactions with the enzyme environment. In addition to a deeper understanding of the racemization mechanism in SerR, based on our simulations we propose specific mutations, which might shift the SerR equilibrium in favor of either l-Ser or d-Ser. Finally, the current studies have produced catalytically competent forms of the rat and human enzymes, which may serve as targets for future docking studies and drug design.

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