Branching Pathways in the Photocycle of Bacteriorhodopsin

The pulsed laser photolysis of light‐adapted bacteriorhodopsin (BR 570 ) is carried out over the temperature range between 25°C and—92°C in neutral and alkaline water‐glycerol solutions. The results arc indicative of considerable complexity, introduced by two temperature dependent branching reactions associated with the intermediates K 610 , L 550 and M 412 , of the BR 570 photocycle. (a) At relatively low temperatures the primary photoproduct K‐ 610 equilibrates with a blue‐shifted species, K p . Both K 610 and the new intermediate subsequently decay into another species, K' r , in a process which competes with the formation of L 550 . Finally, K' p converts very slowly to L 550 . This branched pathway delays the formation of L 550 and thus of M 412 , without affecting the final yield of either species, (b) A thermal back‐reaction regenerating BR 570 takes place at the stage of L 550 , inhibiting the formation of M 412 . The reaction which also predominates at low temperatures, is relatively inefficient at high pH when the forward L 550 → M 412 step is highly catalyzed. It is the superposition of both branching mechanisms, (a) and (b), which accounts for the complex effects of temperature and pH on the photo‐cycle of BR 570 . Mechanism (b) is accounted for by a molecular scheme in which deprotonation of a tyrosine moiety at the stage of L 550 constitutes a prerequisite for deprotonation of the retinal‐lysine schiff‐base as required for forming M 412 . This scheme appears to be directly related to the proton pump. Mechanism (a) introduces additional complexity in the photocycle at low temperatures but its molecular aspects are still unclear.