In vivo characterization of the electrochemical proton gradient generated in darkness in green algae and its kinetic effects on cytochrome b6f turnover

When unicellular algal cells are placed under anaerobic conditions, a large electrochemical gradient is built in darkness across the thylakoid membranes. We have estimated, in vivo, the amplitude of the Delta pH component of this transmembrane potential and shown that the Delta pH is twice as large as the Delta Psi. The amplitude of the Delta<(mu)over tilde>(H+) (similar to 110-140 mV) fits well with estimations based on the ATP/ADP ratio measured in green algae under the same conditions, suggesting that an equilibrium state is established across the thylakoid membrane. Therefore, under anaerobic dark incubation of algae, the electrochemical transmembrane potential is determined only by the cellular ATP content. The existence of this Delta<(mu)over tilde>(H+) is expected to result in a constitutive amount of activated CFo-CF1 ATPase, thereby facilitating ATP synthesis under low light intensity illumination. We report also on the effects of this dark-existing electrochemical gradient on the cytochrome b(6)f complex turnover kinetics. We show that they are largely slowed by the presence of this electrochemical transmembrane potential. The pH component is mainly responsible for the kinetic slowing down of cytochrome b(6)f complex turnover, despite the fact that electrogenicity is associated with the reactions taking place within this complex. Therefore, in vivo, owing to the low lumenal pH, the oxidation of plastoquinol at the Q(o), site is limiting the turnover of the cytochrome b(6)f complex in the presence of the Delta pH, while in its absence the oxidation rate of the bs hemes becomes rate-limiting.