The role of cholesterol in the activity of reconstituted Ca-ATPase vesicles containing unsaturated phosphatidylethanolamine.
The effect of cholesterol on the activity of sarcoplasmic reticulum Ca-ATPase, reconstituted in proteoliposomes containing soybean phosphatidylethanolamine (PE), egg phosphatidylcholine (PC), and cholesterol, was examined. The protein incorporation efficiency increased with PE content but appeared to be independent of cholesterol content. At low cholesterol, PE stimulated calcium uptake. The coupling efficiency of the proteoliposomes increased with an increase in cholesterol content at each PC/(PC + PE) ratio and was more pronounced for those proteoliposomes containing high PE. Dynamic fluorescence measurements of the incorporated lipophilic probe, diphenyl-1,3,5-hexatriene, revealed a decrease in the motion and an increase in the order of the phospholipid fatty acyl chains in proteoliposomes with high cholesterol content. A complementary observation was made using electron spin resonance of the spin label, 2,2-dimethyl-5-dodecyl-5-methyloxazolidine N-oxide. Freeze-fracture electron microscopy studies on proteoliposomes containing 0.20 molar ratio of PC/(PC + PE) and cholesterol revealed predominantly vesicular structures with occasional bilayer defects at high cholesterol content. It is postulated that the cholesterol-induced enhancement of the Ca-transport function of the Ca-ATPase is related to the hydration-related bilayer-destabilizing characteristic of the cholesterol molecule as revealed by 31P NMR.
D000818:Animals; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002784:Cholesterol / Q000494:pharmacology*; D004578:Electron Spin Resonance Spectroscopy; D005227:Fatty Acids / Q000378:metabolism; D005614:Freeze Fracturing; D008081:Liposomes / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008560:Membrane Fluidity / Q000187:drug effects; D008565:Membrane Proteins / Q000378:metabolism; D008854:Microscopy, Electron; D010714:Phosphatidylethanolamines / Q000494:pharmacology*; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013050:Spectrometry, Fluorescence
D005227:Fatty Acids; D008081:Liposomes; D008565:Membrane Proteins; D010714:Phosphatidylethanolamines; D002784:Cholesterol; D000252:Calcium-Transporting ATPases; D002118:Calcium