Fast membrane H+ binding in the light-activated state of Chromatium chromatophores.

Abstract

A fast and sensitive single‐beam spectrophotometer and a variety of flash and steady‐state illumination sources permit the resolution of bromcresol purple absorbancy changes from background absorbancy changes of Chromatium chromatophores, and identification of these fast absorbancy changes with rapid H+ binding by the chromatophores. A saturating light flash gives a rate several hundred times faster than that obtained in continuous light, the maximal rate being about 50 H+/chlorophyll μsec, with a magnitude of the H+: chlorophyll ratio of 1:30 and a duration for the H+ binding reaction, measured by absorbancy changes of bromcresol purple, of 2.2 msec (half‐time, 800 μsec for a 200 μsec flash). A single turnover flash (10 μsec) yields an H+: chlorophyll value of 1:100, with a duration of the H+ change of 800 μsec and a half‐time of 400 μsec. A double‐flash technique shows the recovery for the fast H+ reaction to have a half‐time of 60 msec. The effects of pH, temperature, uncouplers, and inhibitors are also reported.While the hydrogen ion binding would appear to be closely associated with the reduction of an electron acceptor, Y, particularly on the basis of the similar amounts of the two substances (roughly one each per hundred chlorophyll) the reduction time for Y of 27 μsec is over ten‐fold shorter than the H+ binding time in single flash experiments. In addition, the pH sensitivities and activation energies of the two reactions differ.Thus, either an unidentified electron acceptor is involved in the fast H+ changes, or there is no direct relationship between electron transport and H+ binding. If the latter is the case, we propose that electron transport through the photosynthetic center produces in 800 μsec a state of the membrane (a “membrane Bohr effect”) in which hydrogen ion binding is considerably enhanced.