Electron acceptors in isolated intact spinach chloroplasts act hierarchically to prevent over-reduction and competition for electrons

Electron fluxes in isolated intact spinach chloroplasts were analyzed under saturating light and under optimal CO2 and P-i supply. When CO2 assimilation was the only ATP- and NADPH-consuming reaction, the Delta pH decreased and the chloroplasts showed clear evidence of over-reduction. This suggested that additional electron flow is required in order to maintain the Delta pH and the stromal NADPH/ATP ratio. The additional electron flow may be cyclic electron transport around Photosystem I and linear electron transport towards either oxaloacetate or O-2. The contributions of, and the interrelationships between, these three electron transfer pathways were analyzed by following the reactions of chloroplasts in their presence or absence, and by monitoring to what extent they were able to compensate for each other. Inhibition of cyclic electron flow by antimycin A caused strong over-reduction and decreased the Delta pH. Only oxaloacetate, but not O-2, was able to restore photosynthesis. In the presence of H2O2, there was a rapid build-up of a high Delta pH, and the reduction of any other electron acceptor was prevented. It is concluded that the different electron acceptors in the stroma are organized in a hierarchical manner; this allows electron flux towards CO2 and nitrite reduction to proceed without any competition for electrons, and any excess electrons to be taken by these additional non-assimilatory pathways. Hence, the Delta pH is maintained at the required level and over-reduction of the electron transport chain and the stromal redox components is avoided.