Biosynthetic incorporation of fatty acids with photosensitive groups into membrane lipids of cells in tissue culture

Abstract

The physical methods (13C-NMR-spectroscopy and fluorescence spectroscopy) hitherto used for the elucidation of lipid-lipid and lipid-protein interactions in artificial and simple natural membranes were extended to the application of fatty acids, phospholipids and sphingolipids with photochemical labels (azide group) in defined positions, which on photolysis generate nitrenes. These highly reactive groups react with neighbouring molecules, either lipids or polypeptide chains, with insertion or addition. Highly radioactive 12-azido[9,10-3H2]stearic acid, 12-azido[12-3H]oleic acid and 18-azido-[9,10,12,13-3H4]linoleic acid were added to the growth medium of eukaryotic cell lines in tissue culture (BHK 21 cells and Chang liver cells). They were incorporated into neutral, phosphoand sphingolipids in amounts comparable with the unsubstituted parent fatty acids. The distribution of the azido fatty acids in the phospholipids has been determined by enzymatic hydrolysis (phospholipase A2) on the basis of the distribution of their radioactivity. Radio gas chromatography and combined gas chromatography and mass spectroscopy revealed that the azide group of the radioactive fatty acids remained unaltered.