Spin-label study of the relation between enzymatic activity and lipid-protein organization in reconstituted cytochrome c oxidase.
Lipid-depleted cytochrome c oxidase (EC 1.9.3.1) containing less than 20 microgram lipids per milligram protein was reconstituted with pure phospholipids of well-defined chemical structure and fatty acid composition without using detergents and (or) sonication. For the maximal restoration of electron transport activity, lipid-depleted cytochrome c oxidase required acidic phospholipds such as phosphatidylglycerol or phosphatidylserine or lysophospholipids such as lysophosphatidylcholine or lysophosphatidic acid, but no specific phospholipid fatty acid composition was necessary. The organization of the lipid environment of the reconstituted cytochrome c oxidase, having a well-defined lipid composition, morphology, and a high specific activity, was examined by electron spin resonance spectroscopy using 2-(14-carboxytetradecyl)-2-ethyl-4,4-dimethyl-3-oxazolidinyloxyl (16-doxyl stearic acid) and 16-doxyl stearic acid - containing phosphatidylglycerol. The presence of boundary lipid was established in both lamellar and micellar organizations of reconstituted cytochrome c oxidase and was not necessarily related to the enzymatic activity of the complex. Our results have established that aside from structural considerations, the boundary lipid, at least in the reconstituted cytochrome c oxidase, is a necessary but not sufficient condition for the enzymatic expression of cytochrome c oxidase.
D000818:Animals; D002417:Cattle; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D003576:Electron Transport Complex IV / Q000378:metabolism; D008854:Microscopy, Electron; D008929:Mitochondria, Heart / Q000201:enzymology; D010743:Phospholipids / Q000494:pharmacology; D011485:Protein Binding; D011487:Protein Conformation; D013113:Spin Labels
D010743:Phospholipids; D013113:Spin Labels; D003576:Electron Transport Complex IV