Group-specific assays that distinguish between the four major types of mammalian phospholipase A(2)

Phospholipase A(2) (PLA(2)) constitutes a diverse superfamily of enzymes wh ich catalyze the deacylation of phospholipids. At least four types of PLA(2 ) are potentially involved in arachidonic acid release in cells and tissues . Since all of them catalyze the same enzymatic reaction, it is difficult t o distinguish between them in mixtures of enzymes normally present in biolo gical samples. Utilizing specific properties of each PLA(2), we have design ed distinct assay procedures which selectively and sensitively detect each type: Group VI Ca2+-independent PLA(2), Group IV cytosolic Ca2+-dependent P LA(2), Groups V and IIA secreted PLA(2)s. Each specific assay procedure is selective for a particular PLA(2) type by at least fourfold and as high as four orders of magnitude relative to the other three enzymes. All assays ca n detect PLA(2) activity with as low as subnanogram quantities of enzyme, I mportantly, these assays are able to differentiate and quantitate the bioch emically and structurally related enzymes, Group IIA and V sPLA(2)s in crud e biological samples. Employing this system, we have found that iPLA(2) is the dominant PLA(2) in rat brain, and cPLA(2) is the most abundant PLA(2) i n P388D(1) macrophages and human amnionic WISH cells. (C) 1999 Academic Pre ss.