Studies of energy linked reactions: a role for lipoic acid in the purple membrane of Halobacterium halobium.
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Using H. halobium cell envelope vesicles containing either bacteriorhodopsin plus Na+ pump, bacteriorhodopsin alone, Na+ pump alone, or no light-responsive pigment altogether, it could be shown that the large majority of light energized Na+ extrusion in these mutants is linked to bacteriorhodopsin and to protonmotive force, and therefore must be facilitated by a Na+/H+ antiporter. Thus, the recently discovered primary Na+ pump makes only a minor contribution to light-mediated Na+ flux. The activity of the Na+/H+ antiporter appears to be independent of the presence of any photoreactive pigments, since an artifical electron donor will drive rapid Na+ extrusion in all of the vesicle preparations tested.
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The photocycle of the light-adapted purple membrane was studied with a time-resolved fluorometry apparatus: fluorescence of the sample suspension (greater than 660 nm) was pumped with a 633-nm cw laser and the temporal change induced by a 532-nm pulsed laser was measured with a photon-counting-type transient recorder. The formation and the decay of the O640 intermediate were clearly observed in the pH region between 4.0 and 11.4. A photochemical cycle of N560 was apparently driven in alkaline suspension (pH greater than 9.3). An O-like fluorescent intermediate Q appears and decays with time constants of less than 0.1 ms and 1.7 +/- 0.2 ms, respectively.
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