59.
We previously have presented evidence for prominent structural changes in helices F and G of bacteriorhodopsin during the photocycle. These changes were determined by carrying out electron diffraction analysis of illuminated two-dimensional crystals of wild-type bacteriorhodopsin or the Asp-96 → Gly mutant that were trapped at a stage in the photocycle after light-driven proton release, but preceding proton uptake from the aqueous medium. Here, we report structural analysis of the long-lived O intermediate observed in the photocycle of the Leu-93 → Ala mutant, which accumulates after the release and uptake of protons, but before the reisomerization of retinal to its initial all-
trans state. Projection Fourier difference maps show that upon illumination of the Leu-93 → Ala mutant, significant structural changes occur in the vicinity of helices C, B, and G, and to a lesser extent near helix F. Our results suggest that (
i) all four helices that line the proton channel (B, C, F, and G) participate in structural changes during the late stages of the photocycle, and (
ii) completion of the photocycle involves significant conformational changes in addition to those that are associated with steps in proton transport.
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