On the Organization and Mechanism of Prostaglandin Synthetase

Abstract

Abstract The organization and mechanism of prostaglandin synthetase in the vesicular gland of sheep was studied by determining the isotope effects in the transformation of [5,6,8,9,11,12,14,15]-octadeutero-arachidonic acid into prostaglandin E2 (PGE2), 11-dehydro-prostaglandin F2α (PGF2α), PGF2α, and 12-hydroxy-5,8,10-heptadecatrienoic acid. The first two products were formed from deuterated arachidonic acid by cleavage of carbon-deuterium bonds at C-9 and C-11, respectively, whereas the last two products were formed without carbon-deuterium cleavage. The ratio (product ratio) between the first and second category of products increased with time (18 to 300 s) in the presence of glutathione and hydroquinone from 1.8 to 3.5. The ratio between deuterated and unlabeled molecules divided by the same ratio of the substrate (relative D:H ratio) was almost constant during the incubation for PGE2 (0.3 to 0.4) and 11-dehydro-PGF2α (0.6 to 0.7), but increased for PGF2α (from 1.9 to 4.0) and 12-hydroxy-5,8,10-heptadecatrienoic acid in parallel with the product ratio. With l-epinephrine as cofactor, the product ratio remained almost constant during the incubation and also in this case the relative D:H ratio of PFG2α was essentially the same as the product ratio. Elimination of the hydrogen (deuterium) at C-9 in the intermediate endoperoxide is the rate-limiting step in the formation of PGE2 from the endoperoxide. The high relative D:H ratio of PGF2α excludes the existence of a separate synthetase for PGF2α and indicates that a common synthetase provides the endoperoxide precursor for both PGE2 and PGF2α.