Serine racemase catalyzes both the synthesis and the degradation of D-serine, an obligatory co-agonist of the glutamatergic NMDA receptors. It is allosterically controlled by ATP, which increases its activity around 7-fold, with a cooperative binding. Serine racemase has been proposed as a drug target for the treatment of several neuropathologies but, so far, the search has been directed only toward the active site, with the identification of a few, low affinity inhibitors. Following the recent observation that NADH inhibits serine racemase, here we show that the inhibition is partial, with an IC50 of 246±63 mM, several fold higher than NADH intracellular concentrations. At saturating concentrations of NADH, ATP binds with a 2-fold lower affinity and without cooperativity, suggesting ligand competition. NADH also reduces the weak activity of hSR in the absence of ATP, indicating an additional ATP-independent inhibition mechanism. By dissecting the NADH molecule, we discovered that the inhibitory determinant is the N-substituted 1,4-dihydronicotinamide ring. Particularly, the NADH precursor 1,4-dihydronicotinamide mononucleotide exhibited a partial mixed-type inhibition, with a KI of 18 ± 7 mM. Docking simulations suggested that all 1,4-dihydronicotinamide derivatives bind at the interdimeric interface, with the ring positioned in an unoccupied site next to the ATP binding site. This newly recognized allosteric site could be exploited for the design of high affinity serine racemase effectors to finely modulate D-serine homeostasis.
- serine racemase
- allosteric modulation
- ©2016 The Author(s)
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