The allosteric interplay between S-nitrosylation and glycine binding controls the activity of human serine racemase

Human serine racemase (hSR) catalyzes the biosynthesis of D-serine, an obligatory co-agonist of the NMDA receptors. It was previously found that the reversible S-nitrosylation of Cys113 reduces hSR activity. Here, we show by site-directed mutagenesis, fluorescence spectroscopy, mass spectrometry, and molecular dynamics that S-nitrosylation stabilizes an open, less-active conformation of the enzyme. The reaction of hSR with either NO or nitroso donors is conformation-dependent and occurs only in the conformation stabilized by the allosteric effector ATP, in which the epsilon-amino group of Lys114 acts as a base toward the thiol group of Cys113. In the closed conformation stabilized by glycine-an active-site ligand of hSR-the side chain of Lys114 moves away from that of Cys113, while the carboxyl side-chain group of Asp318 moves significantly closer, increasing the thiol pK(a) and preventing the reaction. We conclude that ATP binding, glycine binding, and S-nitrosylation constitute a three-way regulation mechanism for the tight control of hSR activity. We also show that Cys113 undergoes H2O2-mediated oxidation, with loss of enzyme activity, a reaction also dependent on hSR conformation.

Automated summary

The reversible S-nitrosylation of human serine racemase (hSR) was found to stabilize an open, less-active conformation of the enzyme, affecting its activity. It was also shown that a three-way regulation mechanism involving ATP binding, glycine binding, and S-nitrosylation tightly controls hSR activity. Additionally, Cys113 undergoes H2O2-mediated oxidation, with loss of enzyme activity, depending on hSR conformation.