Regional distribution, ontogeny, purification, and characterization of theCa2+-independent phospholipase A(2) from rat brain

We purified an 80-kDa Ca2+-independent phospholipase A(2) (iPLA(2)) from ra t brain using octyl-Sepharose, ATP-agarose, and calmodulin-agarose column c hromatography steps. This procedure gave a 30,000-fold purification and yie lded 4 mu g of a near-homogeneous iPLA(2) with a specific activity of 4.3 m u mol/min/mg. Peptide sequences of the rat brain iPLA(2) display considerab le homology to sequences of the iPLA(2) from P388D, macrophages, Chinese ha mster ovary cells, and human B lymphocytes. Under optimal conditions, the i PLA(2) revealed the following substrate preference toward the fatty acid ch ain in the sn-2 position of phosphatidylcholine: linoleoyl > palmitoyl > ol eoyl > arachidonoyl. The rat brain iPLA(2) also showed a head group prefere nce for choline greater than or equal to ethanolamine much greater than ino sitol. The iPLA(2) is inactivated when exposed to pure phospholipid vesicle s. The only exception is vesicles composed of phosphatidylcholine and phosp hatidylinositol 4,5-bisphosphate. Studies on the regional distribution and ontogeny of various phospholipase A(2) (PLA(2)) types in rat brain indicate that the IPLA(2), is the dominant PLA(2) activity in the cytosolic fractio n, whereas the group IIA secreted PLA(2) is the dominant activity in the pa rticulate fraction. The activities of these two enzymes change during postn atal development.