MUTATION OF SERINE-516 IN HUMAN PROSTAGLANDIN G H SYNTHASE-2 TO METHIONINE OR ASPIRIN ACETYLATION OF THIS RESIDUE STIMULATES 15-R-HETE SYNTHESIS/

Prostaglandin G/H synthase (PGHS) is a key enzyme in cellular prostagl andin (PG) synthesis and is the target of non-steroidal anti-inflammat ory agents. PGHS occurs in two isoforms, termed PGHS-1 and PGHS-2 Thes e isoforms differ in several respects, including their enzymatic activ ity following acetylation by aspirin. While PG synthesis by both isofo rms is inhibited by aspirin, 15-R-hydroxyeicosatetraenoic acid (15-R-H ETE) synthesis by PGHS-2, but not PGHS-1, is stimulated by preincubati on with aspirin. We have mutated the putative aspirin acetylation site of hPGHS-2, and expressed the mutants in COS-7 cells using recombinan t vaccinia virus. Enzyme activity and inhibitor sensitivity studies pr ovide evidence that Ser(516) is the aspirin acetylation site of human PGHS-2 and that substitution of a methionine residue at this position can mimic the effects of aspirin acetylation on enzyme activity.