Knowledge Bank

Kinetic resonance Raman spectroscopy $techniques1$ were employed to study the photocycle of the bacteriorhodopsin from the purple membrane of H. halobium. A suspension of purple membrane fragments in $H2O$ and $D2O$ at different pH values was flowed at variable speeds through a laser beam which triggers the photochemical event and is also used to excite the Raman scattering of the intermediate species produced. These studies indicate that there is a hitherto unknown intermediate, preceding the $M412$ species. In these two species the Schiff-base linkage between the retinal chromophore and an $\in$-amino group of a lysine from the protein, is unprotonated, whereas in all the earlier intermediates this linkage is protonated. By tracing the intensity of Raman bands assigned to a protonated and unprotonated $Schiff-basea,2$ we have shown that the rate of deprotonation is Increased at higher pH and exhibits a pH value of 10.2. This suggests that a second lysin from the protein interacts with the Schiff base linkage. Based on this and other results we propose a molecular mechanism for the proton pumping across the membrane which occurs during the photo-cycle of bacteriorhodopsin.