Use of the fluorescent pH probe pyranine to detect heterogeneous directions of proton movement in bacteriorhodopsin reconstituted large liposomes

AbstractLight-induced proton pumping was measured in bacteriorhodopsin reconstituted large proteoliposomes (prepared by reverse-phase evaporation) using the fluorescence of internally trapped pyranine. From the effect of the one-sided inhibitor La3+, the presence of both inside-out and right-side-out oriented bacterio-rhodopsin was demonstrated. The use of pyranine at acidic pH allowed us to detect two distinct populations of proteoliposomes which exhibited oppositely directed proton pumping. This suggests that the two orientations, of bacteriorhodopsin are at least partially distributed in different proteoliposomes