High-pressure extraction of membrane-associated protein kinase C from rat brain.
Extraction of rat brain membrane-associated protein kinase C with high specific activity was obtained by applying benzyl alcohol (a membrane fluidizer), EDTA, and high hydrostatic pressures. Approximately 50% of total brain-associated activity was extracted from membranes. The pressure-extracted activity had an eightfold enrichment in the lipid/protein ratio when compared with the cytosolic fraction. This may explain the inability of exogenous diacylglycerol to stimulate endogenous phosphorylation in pressure-extracted activity. The enzyme is extracted at greater than 1,300 atm, a result indicating it most likely has a portion inserted into the hydrophobic portion of the membrane bilayer. Perturbation of the native membrane induces a change in the membrane-associated protein kinase C-lipid interaction that permits extraction under conditions used for the cytosolic species. This is the first report of conversion of the endogenous membrane species to a cytosolic one and may be important in determining the role of protein kinase C in neuronal regulation.
Lester
D S
DS
Department of Membrane Research, Weizmann Institute of Science, Rehovot, Israel.
D000818:Animals; D001921:Brain / Q000201:enzymology*; D003600:Cytosol / Q000201:enzymology; D006874:Hydrostatic Pressure; D008297:Male; D008566:Membranes / Q000201:enzymology; D009448:Neurochemistry / Q000379:methods*; D011312:Pressure; D011493:Protein Kinase C / Q000032:analysis*; D051381:Rats; D011919:Rats, Inbred Strains; D013347:Subcellular Fractions / Q000201:enzymology
D011493:Protein Kinase C