Studies on well-coupled Photosystem-I-enriched subchloroplast vesicles. Kinetic aspects of flash-induced energy transduction

Abstract

Flash-induced ATP synthesis, coupled to cyclic electron flow in Photosystem-I-enriched subchloroplast vesicles (from spinach) was studied by continuous registration of luciferin-luciferase luminescence. In flash trains, the ATP yield per flash was in any case constant after 2–3 flashes and remained so throughout the train. However, valinomycin (not nigericin) induced a lag phase which lasted about 10 flashes at 0.1–0.5 Hz and about 50 flashes at 5 Hz. Valinomycin (10–100 nM) as well as nigericin (35–100 nM) or the uncoupler FCCP (50–250 nM) partly inhibited phosphorylation throughout the flash train; a combination of 50 nM of each inhibited phosphorylation completely. In the absence of ionophores, the flash-induced energization increase seems to be mainly dependent on electric-potential changes, while a pH gradient (ΔpH) only contributes to the steady-state energization level in the dark. In trains of single flashes, the ATP yield increased with flash frequency, but only between about 0.1 and 0.8 Hz. A model is presented, which explains this behaviour assuming a sharp, non-linear increase of the turnover rate of the ATPase molecules above a certain level of the proton-motive force (Δ\\̃gmH+. In groups containing two or three flashes at variable intervals, the ATP yield per flash was sharply decreased at intervals within the group shorter than 40–50 ms, especially in the presence of nigericin or the uncoupler FCCP. The decrease seemed to be less pronounced in the presence of electron-transfer inhibitors. It is probably due to kinetic limitation of electron transfer and proton translocation. Nigericin and FCCP have a larger effect on single flashes than on flash groups. Their effect supports the assumption that the ATPase turnover rate depends non-linearly on Δ\\̃gmH+.