REORIENTATIONS IN THE BACTERIORHODOPSIN PHOTOCYCLE

Reversible photoinduced reorientations of bacteriorhodopsin have been detected in suspensions of the purple membrane of Halobacterium salina rium. The anisotropy in bacteriorhodopsin during the nanosecond throug h millisecond stages of the photocycle was measured by time-resolved l inear dichroism and transient absorption measurements. From these meas urements the anisotropies of the K, L, M, and O intermediates were det ermined and related to the chromophore orientation with respect to the initially selected orientation. The anisotropies of the K and L state s are 0.38 +/- 0.01 and 0.35 +/- 0.01, respectively. Further anisotrop y decay after formation of the M intermediate in about 0.5 ms is evide nce of orientational motion at this stage in the photocycle. A constan t anisotropy with a value of 0.39 +/- 0.02 in the O intermediate demon strates a recovery of the initial protein orientation with the formati on of the O state. These results demonstrate that reorientations in BR are photoinduced and reversible. Similar measurements for L and M wer e carried out for purple membrane in polyacrylamide gels, where the an isotropies in the L and M states are 0.38 +/- 0.014 and 0.36 +/- 0.01, respectively. These results show that reorientations also occur in BR immobilized in gels. Anisotropy decay in the M state after formation of the M intermediate was not detected in the gels, in contrast to the M intermediate in suspensions. Orientational changes are observed for BR in purple membrane suspensions in the K state, during the K --> L step, in the M state possibly related to an M(1) --> M(2) transition, and in the O state, where an almost complete return to the original or ientation occurs. For BR in gels, reorientations are reduced but not a bolished. The possible role of these reorientations in bacteriorhodops in is discussed.