Lipid vesicles which can bind to protein kinase C and activate the enzyme in the presence of EGTA.
Maximal protein kinase C activity with vesicles of phosphatidic acid and 1,2-dioleoyl-sn-glycerol is observed in the absence of added Ca2+. Addition of phosphatidylcholine to these vesicles restores some calcium dependence of enzyme activity. 1,2-Dioleoyl-sn-glycerol eliminates the Ca(2+)-dependence of protein kinase C activity found with phosphatidic acid alone. Phorbol esters do not mimic the action of 1,2-dioleoyl-sn-glycerol in this respect. This suggests that the 1,2-dioleoyl-sn-glycerol effect is a result of changes it causes in the physical properties of the membrane rather than to specific binding to the enzyme. The effect of 1,2-dioleoyl-sn-glycerol on the phosphatidic-acid-stimulated protein kinase C activity is dependent on the molar fraction of 1,2-dioleoyl-sn-glycerol used and results in a gradual shift from Ca2+ stimulation at low 1,2-dioleoyl-sn-glycerol concentrations to calcium inhibition at higher concentrations of 1,2-dioleoyl-sn-glycerol. Phosphatidylserine-stimulated activity is also shown to be largely independent of the calcium concentration at higher molar fractions of 1,2-dioleoyl-sn-glycerol. Thus, with certain lipid compositions, protein kinase C activity becomes independent of the calcium concentration or requires only very low, stoichiometric binding of Ca2+ to high affinity sites on the enzyme. Protein kinase C can bind to phosphatidic acid vesicles more readily than it can bind to phosphatidylserine vesicles in the absence of calcium. Addition of 1,2-dioleoyl-sn-glycerol to phosphatidylserine vesicles promotes the partitioning of protein kinase C into the membrane in the absence of added Ca2+. There is no isozyme specificity in this binding. These results suggest that a less-tightly packed headgroup region of the bilayer causes increased insertion of protein kinase C into the membrane. This is a necessary but not sufficient condition for activation of the enzyme in the presence of EGTA.
Epand
R M
RM
Department of Biochemistry, McMaster University, Ontario, Canada.
Stafford
A R
AR
Lester
D S
DS
D000818:Animals; D001921:Brain / Q000201:enzymology; D002118:Calcium / Q000494:pharmacology; D004075:Diglycerides / Q000378:metabolism / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D015151:Immunoblotting; D007527:Isoenzymes / Q000378:metabolism; D008051:Lipid Bilayers / Q000378:metabolism; D008081:Liposomes / Q000378:metabolism*; D010712:Phosphatidic Acids / Q000378:metabolism / Q000494:pharmacology; D010713:Phosphatidylcholines / Q000378:metabolism / Q000494:pharmacology; D010718:Phosphatidylserines / Q000378:metabolism / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats
D004075:Diglycerides; D007527:Isoenzymes; D008051:Lipid Bilayers; D008081:Liposomes; D010712:Phosphatidic Acids; D010713:Phosphatidylcholines; D010718:Phosphatidylserines; D004533:Egtazic Acid; D011493:Protein Kinase C; D002118:Calcium; C013965:diolein