Partial purification and characterization of a (glycosyl) inositol phospholipid-specific phospholipase C from peanut.
We have isolated a glycosyl inositol phospholipid (GIP) anchor-hydrolyzing activity from peanut seeds by a series of column chromatographic steps. The activity has a pH optimum below 6.0, requires calcium, and is inhibited by sulfhydryl reagents. It cleaves the GIP anchors of solubilized acetylcholinesterase from bovine erythrocytes and variant surface glycoprotein from Trypanosoma brucei. On the other hand, it does not act on membrane-bound GIP-anchored substrate or on inositol-acylated GIP anchor of human erythrocyte acetylcholinesterase. The only product released from [3H]myristate-labeled variant surface glycoprotein following treatment with the activity from peanut was 3H-labeled diacylglycerol. Together, these findings identify the activity from peanut seeds as a GIP anchor-hydrolyzing phospholipase C. The enzyme has been found to hydrolyze not only protein GIP anchors but also phosphatidylinositol, whereas it shows no activity against other phospholipids. The water-soluble products of phosphatidylinositol hydrolysis by peanut phospholipase C were characterized as a mixture of inositol 1,2-cyclic phosphate and inositol phosphate.
Bütikofer
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P
Institute of Biochemistry & Molecular Biology, University of Bern, Switzerland.
Brodbeck
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U
D000110:Acetylcholinesterase / Q000097:blood; D010367:Arachis / Q000201:enzymology*; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D002852:Chromatography, Ion Exchange; D004912:Erythrocytes / Q000201:enzymology; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D007295:Inositol Phosphates / Q000378:metabolism; D007700:Kinetics; D010718:Phosphatidylserines / Q000378:metabolism; D013379:Substrate Specificity; D010738:Type C Phospholipases / Q000302:isolation & purification / Q000378:metabolism*
D017261:Glycosylphosphatidylinositols; D007295:Inositol Phosphates; D010718:Phosphatidylserines; D000110:Acetylcholinesterase; D010738:Type C Phospholipases