Temperature dependence of light-induced proton movement in reconstituted purple membrane.
Bacteriorhodopsin (BR) was incorporated into phosphatidylcholine (PC) vesicles containing different amounts of other lipids. Under the conditions of nullified membrane potential, light-induced proton movement seemed to follow a kinetic scheme which assumed the existence of a proton-pumping inhibition process characterized by a rate constant, kI. The temperature dependence of both kI and the membrane proton leak rate constant (kD) obeyed a simple Arrhenius equation. The presence of cholesterol in the membrane significantly increased the activation energy (Ea) of both the inhibition and leak process. However, further addition of phosphatidic acid (PA) suppressed the increase of Ea associated with kI. The initial proton pumping rate (R0) of vesicles reconstituted with PC showed a bell-shaped temperature dependence with a maximum at approximately 20 degrees C. The addition of cholesterol abolished this dependence. These results suggest that the molecular origin of the inhibition process characterized by kI is different from that of R0 or kD. The temperature dependence of the steady-state fluorescence polarization of dansylated bacteriorhodopsin in vesicles was also investigated. The polarization of the labels in the vesicles without cholesterol showed a bell-shaped temperature dependence with a maximum at approximately 20 degrees C. However, in the presence of cholesterol, the polarization increased linearly as temperature decreased. A comparison of these results with the observed proton movement in similarly reconstituted systems with unmodified protein indicates that membranes with a low fluidity and negatively charged surfaces enhance proton pumping efficiency of bacteriorhodopsin.
D001436:Bacteriorhodopsins / Q000378:metabolism* / Q000528:radiation effects; D001693:Biological Transport, Active / Q000528:radiation effects; D002338:Carotenoids / Q000378:metabolism*; D005454:Fluorescence Polarization; D006217:Halobacterium; D007700:Kinetics; D008027:Light; D010743:Phospholipids / Q000378:metabolism; D011522:Protons; D013696:Temperature
D010743:Phospholipids; D011522:Protons; D002338:Carotenoids; D001436:Bacteriorhodopsins