| Bacteriorhodopsin (BR) functions as a light-driven proton pump in Purple Membrane (PM). Upon the absorption of light, BR goes through a photochemical cycle comprised of K, L, M, N, O five distinct states. The M state and its decay kinetics are considered important to the proton release of BR.In this paper, we reconstituted BR into proteoliposome with different formulation and various proportion of charged lipid, and studied the activity of BR in helping proton cross membrane, the effect of charged lipid on reconstitution mechanism of BR as well as its photochemical cycle activity.For the proteoliposome with neutral lipid, BR could help protons cross the membrane effectively under no illumination. Reasons are hypothesized to be: (1) BR could act as a proton channel; (2) the insertion of BR into lipid membrane raises its permeability. We also found that when raising the percentage of anionic lipid proportion, the effect of BR was reduced and even disappeared when it reached 20%; in terms of the cationic liposome, BR's effect was also reduced, while the permeability of proteoliposome only changed little when changing the percentage of cationic lipid from 10% to 30%.Extrinsic charged lipid could affect the insertion direction of BR during reconstitution; the proteoliposome with more anionic lipid had a higher proportion of BR in inside-in direction. However, we couldn't get the proteoliposome with BR all in inside-in direction even in pure anionic lipid environment. We agreed that charge is a main, but not the only, factor that could affect the insertion mechanism of BR.The M decay kinetic of BR varied with the proportion of the charged lipid in proteoliposome. With the increase of the percentage of cationic lipid (0～60%), the decay time of BR prolonged, and anionic lipid component could decrease the decay life of M state. We supposed that the rearrangement of charge inside BR induced by the change of ion concentration in micro-environment could affect the photochemical cycle of BR.
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