Bacteriorhodopsin: lipid environment and conformational changes.

The polar lipids of the purple membrane were exchanged for different phosphatidylcholine species. The resulting complexes had the same protein to lipid-phosphorus ratio as the natural membrane, but only about 0.5-1.0 mole of original lipid was still present per mole of bacteriorhodopsin. In such complexes the bacteriorhodopsin photocycle is slowed down 10-20 times, but the strong protein-protein interaction is not abolished. Due to the slow rate of the photocycle we were able to measure in the light the ratio between net proton release and net accumulation of the last intermediate of the photocycle, the unprotonated M412. This ratio was not constant and equal to 1.0, as expected for a single deprotonation reaction, but varied with pH from 1.5 to 0.4. The variable ratio suggests that light-induced conformational changes occur in the nonchromophore part of the protein, which shift the pKa values of unidentified groups so as to cause binding or release of additional protons. A similar conclusion was drawn from experiments on the kinetics of proton transfer by bacteriorhodopsin in subbacterial particles of Halobacterium halobium and in reconstituted bacteriorhodopsin proteoliposomes. However, in this case light-induced association and dissociation of additional protons occurs simultaneously on different sides of the membrane.