NOVEL PHOSPHOSERINE PHOSPHATASE INHIBITORS

Phosphoserine phosphatase (EC 3.1.1.3) catalyzes the final step in the major pathway of L-serine biosynthesis in brain. This enzyme may also regulate the levels of glycine and D-serine, the known and putative c o-agonists fur the glycine site of the N-methyl-D-aspartate receptor i n caudal and rostral brain regions, respectively. Using L-phosphoserin e as substrate, the rank order potency for inhibition of phosphoserine phosphatase was p-chloromercuriphenylsulfonic acid (CMPSA) > glycerop hosphorylcholine much greater than hexadecylphosphocholine greater tha n or equal to phosphorylcholine > N-ethylmaleimide greater than or equ al to L-serine > fluoride > D-2-amino-3-phosphonopropionic acid (D-AP3 ). Glycerylphosphorylcholine (IC50 18 mu M) was found to be an uncompe titive inhibitor of phosphoserine phosphatase. Glycerylphosphorylcholi ne probably binds a novel site on the enzyme since the known allosteri c inhibitor L-serine is highly selective for its feedback regulatory s ite, indicated by the inactivity of 25 L-serine analogs. Fluoride ion (IC50 770 mu M) may bind the active site as has been shown for other M g2+-dependent enzymes. The sulfhydryl reagent CMPSA is a potent, nonco mpetitive inhibitor of the enzyme using L-phosphoserine as substrate ( IC50 9 mu M) but is > 300-fold less potent using D-phosphoserine as su bstrate. Substrate-dependent differences are also observed with the su lfhydryl alkylator N-ethylmaleimide, which inhibits L-phosphoserine, b ut stimulates D-phosphoserine hydrolysis. These sulfhydryl reagents ma y dissociate multimeric forms of the enzyme to form monomers; the mult imeric forms and monomers may preferentially cleave L- and D-phosphose rine, respectively. Phosphorylcholine esters and sulfhydryl reagents m ay prove useful in determining the contribution of phosphoserine phosp hatase to the biosynthesis of glycine and D-serine in neuronal tissue in vitro. (C) 1997 Elsevier Science B.V.