Proton translocation in intact cells of the photosynthetic bacterium Rhodopseudomonas capsulata

(1) Cresol red and phenol red are found to be rapid and reliable indicators of external pH in intact cell suspensions of Rhodopseudomonas capsulata . (2) Light-induced pH changes measured with the indicators in 10 mM Na 2 SO 4 are complex. A rapid acidification (phase A, t 1 2 ≈ 30 ms , 1 H + per 29 bacteriochlorophyll) was followed by a lag (phase B, approx. 300 ms duration) and then an extensive and slow acidification (phase C, duration, approx. 2 s, 1 H + per 0.8 bacteriochlorophyll). The pH reached a minimum after 2.5 s (phase D) and subsequently began to increase (phase E). (3) Phase A appears to be a component of outwardly directed, electrogenic proton translocation coupled directly to electron transport. The effects of antimycin A, venturicidin and an uncoupling agent, and the kinetics of the change during and after single and double light pulses, are fully consistent with this view. (4) The origin of the apparent lag, phase B is unknown, but it may be unrelated to chemiosmotic proton translocation. Events occurring during phase B distort observations on the kinetics of the proton-translocating activity of the bacterial membrane when recorded on a longer time scale. (5) Phase C does not coincide with a membrane-potential-generating component (judged by the kinetics of the electrochromic band shift) and is probably the result of electron-transport-driven, electrically neutral H + efflux accompanying the efflux of anions from, or uptake of cations by the cell. Because it is so extensive, phase C dominates the response of a glass electrode to pH during the first few seconds of illumination. (6) Observations on H + efflux measured with glass electrodes following respiratory pulses in Escherichia coli (Gould, J.M. and Cramer, W.A. (1977) J. Biol. Chem. 252, 5875–5882) and in Paracoccus denitrificans (Hitchens, G.D. and Kell, D.B. (1984) Biochim. Biophys. Acta 766, 222–232) which were thought to be inconsistent with the chemiosmotic coupling hypothesis, are confirmed in Rhodopseudomonas capsulata . These observations are discussed with reference to the processes observed on a more rapid time-scale with pH indicators.