Regional distribution, ontogeny, purification, and characterization of the Ca2+-independent phospholipase A2 from rat brain.
We purified an 80-kDa Ca2+-independent phospholipase A2 (iPLA2) 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 microg of a near-homogeneous iPLA2 with a specific activity of 4.3 micromol/min/mg. Peptide sequences of the rat brain iPLA2 display considerable homology to sequences of the iPLA2 from P388D1 macrophages, Chinese hamster ovary cells, and human B lymphocytes. Under optimal conditions, the iPLA2 revealed the following substrate preference toward the fatty acid chain in the sn-2 position of phosphatidylcholine: linoleoyl > palmitoyl > oleoyl > arachidonoyl. The rat brain iPLA2 also showed a head group preference for choline > or = ethanolamine >> inositol. The iPLA2 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 A2 (PLA2) types in rat brain indicate that the iPLA2 is the dominant PLA2 activity in the cytosolic fraction, whereas the group IIA secreted PLA2 is the dominant activity in the particulate fraction. The activities of these two enzymes change during postnatal development.
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Department of Chemistry and Biochemistry, Revelle College and School of Medicine, University of California, San Diego, La Jolla 92093-0601, USA.
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Dennis
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D000818:Animals; D001921:Brain / Q000033:anatomy & histology / Q000201:enzymology*; D002118:Calcium / Q000502:physiology*; D006224:Cricetinae; D006868:Hydrolysis; D007527:Isoenzymes / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D008823:Micelles; D017830:Octoxynol; D010741:Phospholipases A / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D054467:Phospholipases A2; D051381:Rats; D017207:Rats, Sprague-Dawley; D016415:Sequence Alignment; D013379:Substrate Specificity; D014357:Trypsin
D007527:Isoenzymes; D008823:Micelles; D017830:Octoxynol; D010741:Phospholipases A; D054467:Phospholipases A2; D014357:Trypsin; D002118:Calcium