In vivo analysis of the effect of dicyclohexylcarbodiimide on electron and proton transfers in cytochrome bf complex of Chlorella sorokiniana

The effect of N,N'-dicyclohexylcarbodiimide (DCCD) on electron and proton transfers within the cytochrome (cyt) bf complex has been analyzed in living cells of the green algae Chlorella sorokiniana under anaerobic conditions. DCCD induces a partial decoupling of the protomotive Q-cycle, in agreement with the conclusions of Wang and Beattie (1991) Arch. Biochem. Biophys. 291, 363-370. In the presence of 20 mu M DCCD, we observe the development of a lag phase in the kinetics of the slow electrogenic phase associated with electron and proton transfers within the cyt bf complex. In the same conditions, the initial rate of cyt b and cyt f reduction is decreased by about 30%. We propose that in the absence of DCCD, a transmembrane movement of proton is coupled to the oxidation of plastoquinol at site Q(o),. In the presence of 20 mu M DCCD, this redox-coupled proton pump is inhibited, and the kinetics of phase b and cyt b reduction become close to that predicted on the basis of a pure Q-cycle process. In agreement with this hypothesis, we observe that upon a weak-flash excitation, two charges are translocated through the membrane in addition to the charge translocated at the level of photosystem I. Part of this large electrogenic phase could be associated with the translocation of a proton from the stroma to the lumen. A tentative mechanism is discussed that remains in the frame of the Q-cycle but accounts for an additional proton-pumping process or for the partial decoupling observed in the presence of DCCD, as well.