Group IV cytosolic phospholipase A2 binds with high affinity and specificity to phosphatidylinositol 4,5-bisphosphate resulting in dramatic increases in activity.
The group IV cytosolic phospholipase A2 (cPLA2) exhibits a potent and specific increase in affinity for lipid surfaces containing phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) at physiologically relevant concentrations. Specifically, the presence of 1 mol% PtdIns(4,5)P2 in phosphatidylcholine vesicles results in a 20-fold increase in the binding affinity of cPLA2. This increased affinity is accompanied by an increase in substrate hydrolysis of a similar magnitude. The binding studies and kinetic analysis indicate that PtdIns(4,5)P2 binds to cPLA2 in a 1:1 stoichiometry. The magnitude of the effect of PtdIns(4,5)P2 is unique among anionic phospholipids and larger than that for other polyphosphate phosphatidylinositols. The effect of PtdIns(4,5)P2 on the activity of cPLA2 is at least an order of magnitude larger than the concomitant changes in the fraction of the enzyme associated with lipid membranes. Striking parallels between the interaction of cPLA2 with PtdIns(4,5)P2 and the interaction of the pleckstrin homology domain of phospholipase C delta 1 with PtdIns(4,5)2 combined with sequence analysis of cPLA2 lead us to propose the existence and location of a pleckstrin homology domain in cPLA2. We further show that the very nature of the interaction of proteins such as cPLA2 with multiple ligands incorporated into membranes follows a specific model which necessitates the use of an experimental methodology suitable for a membrane interface to allow for a meaningful analysis of the data.
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Department of Chemistry and Biochemistry, Revelle College, University of California, San Diego, La Jolla 92093-0601, USA.
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Dennis
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D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001798:Blood Proteins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008081:Liposomes / Q000378:metabolism; D008823:Micelles; D008969:Molecular Sequence Data; D010714:Phosphatidylethanolamines / Q000378:metabolism; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism*; D010718:Phosphatidylserines / Q000378:metabolism; D054803:Phospholipase C delta; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D010750:Phosphoproteins; D011817:Rabbits; D011994:Recombinant Proteins / Q000378:metabolism; D013049:Spectrin / Q000737:chemistry / Q000378:metabolism; D010738:Type C Phospholipases / Q000737:chemistry / Q000378:metabolism
D001798:Blood Proteins; D007527:Isoenzymes; D008081:Liposomes; D008823:Micelles; D010714:Phosphatidylethanolamines; D019269:Phosphatidylinositol 4,5-Diphosphate; D010718:Phosphatidylserines; D010750:Phosphoproteins; D011994:Recombinant Proteins; C039077:platelet protein P47; D013049:Spectrin; D010741:Phospholipases A; D054467:Phospholipases A2; D010738:Type C Phospholipases; C518024:PLCD1 protein, human; D054803:Phospholipase C delta