The mechanism of H+ transfer by bacteriorhodopsin The properties and the function of intermediate P

Abstract
Regeneration of bacteriorhodopsin (bR) from the M intermediate via a new intermediate P has been studied. In the purple sheets treated with 0.015% Triton X‐100 (the P → bR transition is suppressed), the α‐maximum of absorption of P is located at 560 ± 5 nm, the extinction coefficient being equal to 0.7 ± 0.1 of that of bR. Besides, there is a small but measurable absorbance increase at 330–350 nm, which seems to be due to a β‐maximum of 13‐cis‐retinal residue in P. The similarity of the α‐maximum of P and bR suggests the Schiff base nitrogen to be protonated at the M → P transition. The kinetics of P → bR transition measured at the 335 nm absorbance decrease coincides with that of proton uptake accompanying bR regeneration after flash. A scheme is proposed assuming that H+ absorption from the water phase is coupled to the 13‐cis → all‐trans isomerization of retinal residue in which the Schiff base is already protonated by a proton‐donor group of the protein.