Regional distribution, ontogeny, purification, and characterization of the Ca2+-independent phospholipase A2 from rat brain

We purified an 80-kDa Ca2+-independent phospholipase A(2) (iPLA(2)) from rat brain using octyl-Sepharose, ATP-agarose, and calmodulin-agarose column chromatography steps. This procedure gave a 30,000-fold purification and yielded 4 mu g of a near-homogeneous iPLA(2) with a specific activity of 4.3 mu mol/min/mg. Peptide sequences of the rat brain iPLA(2) display considerable homology to sequences of the iPLA(2) from P388D, macrophages, Chinese hamster ovary cells, and human B lymphocytes. Under optimal conditions, the iPLA(2) revealed the following substrate preference toward the fatty acid chain in the sn-2 position of phosphatidylcholine: linoleoyl > palmitoyl > oleoyl > arachidonoyl. The rat brain iPLA(2) also showed a head group preference for choline greater than or equal to ethanolamine much greater than inositol. The iPLA(2) is inactivated when exposed to pure phospholipid vesicles. The only exception is vesicles composed of phosphatidylcholine and phosphatidylinositol 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 fraction, whereas the group IIA secreted PLA(2) is the dominant activity in the particulate fraction. The activities of these two enzymes change during postnatal development.