31P-nuclear magnetic resonance and freeze-fracture electron microscopic studies on reconstituted bacteriorhodopsin vesicles

Bacteriorhodopsin has been reconstituted into egg-phosphatidylcholine vesicles by various methods. The resulting preparations have been analyzed on density gradients and by freeze-fracture electron microscopy. The homogeneity of the vesicle preparations and the light-induced intravesicular pH changes have been studied by 31P-NMR, using glucose 6-phosphate as pH probe. It is concluded that bacteriorhodopsin is incorporated in the inside-out mode in vesicles up to about 100 nm. Above this diameter, more or less random insertion takes place.