Interfacial activation, lysophospholipase and transacylase activity of group VI Ca2+-independent phospholipase A2.
The Group VI 80-kDa Ca2+-independent phospholipase A2 (iPLA2) has been purified from murine P388D1 macrophages and Chinese hamster ovary (CHO) cells. The amino acid sequence of the iPLA2 has been determined and shown to contain a lipase consensus sequence and eight ankyrin repeats, which makes it distinct from Group I-V PLA2s. This enzyme appears to play a key role in mediating basal phospholipid remodeling. We now report that the Group VI iPLA2 displays interfacial activation toward short chain phospholipids, 1-octanoyl-2-heptanoyl-sn-glycero-3-phosphocholine, 1,2-diheptanoyl-sn-glycero-3-phosphocholine, and 1,2-dihexanoyl-sn-glycero-3-phosphocholine micelles. ATP protects the iPLA2 from a loss in activity as a result of prolonged incubation during the assay. Hence higher enzyme activity is observed in the presence than in the absence of ATP. Similar protection was obtained with glycerol. In addition, the iPLA2 exhibits multiple activities which are strongly dependent on substrate presentation. The lysophospholipase activity of this enzyme was diminished by Triton X-100 and stimulated by glycerol. With the combination of 50 microM Triton X-100 and 50% glycerol, the enzyme's lysophospholipase activity achieved equivalent activity to its PLA2 activity. The iPLA2 displayed both lysophospholipid/transacylase and phospholipid/transacylase activity, supporting the conclusion that the mechanism of action of iPLA2 proceeds through an acyl-enzyme intermediate as proposed for the Group IV cPLA2.
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Department of Chemistry and Biochemistry, University of California at San Diego, La Jolla, CA 92093-0601, USA.
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D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase; D000217:Acyltransferases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000494:pharmacology*; D006224:Cricetinae; D004789:Enzyme Activation; D005990:Glycerol / Q000494:pharmacology; D007700:Kinetics; D008049:Lipase / Q000737:chemistry; D008245:Lysophospholipase / Q000378:metabolism*; D008264:Macrophages / Q000201:enzymology; D051379:Mice; D009097:Multienzyme Complexes / Q000378:metabolism*; D010741:Phospholipases A / Q000737:chemistry / Q000378:metabolism*; D054467:Phospholipases A2; D010743:Phospholipids / Q000494:pharmacology
D009097:Multienzyme Complexes; D010743:Phospholipids; D000255:Adenosine Triphosphate; D000217:Acyltransferases; D008049:Lipase; D010741:Phospholipases A; D054467:Phospholipases A2; D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase; D008245:Lysophospholipase; C021782:lysophospholipase-transacylase; D005990:Glycerol; D002118:Calcium