Quantitation of plastoquinone photoreduction in spinach chloroplasts

The question of plastoquinone (PQ) concentration and its stoichiometry to photosystem I (PSI) and PSII in spinach chloroplasts is addressed here. The results from three different experimental approaches were compared. (a) Quantitation from the light-induced absorbance change at 263 nm (ΔA263) yielded the following ratios (mol:mol); Chl:PQ=70:1, PQ:PSI=9:1 and PQ:PSIIα=7:1. The kinetics of PQ photoreduction were a monophasic but non-exponential function of time. The deviation of the semilogarithmic plots from linearity reflects the cooperativity of several electron transport chains at the PQ pool level. (b) Estimates from the area over the fluorescence induction curve (Afl) tend to exaggerate the PQ pool size because of electron transfer via PSI to molecular oxygen (Mehler reaction) resulting in the apparent increase of the pool of electron acceptors. The reliability of the Afl method is increased substantially upon plastocyanin inhibition by KCN. (c) Quantitation of the number of electrons removed from PQH2 by PSI, either under far-red excitation or after the addition of DCMU to preilluminated chloroplasts, is complicated due to the competitive loss of electrons from PQH2 to molecular oxygen. The latter is biphasic reaction occurring with half-times of about 2 s (30-40% of PQH2) and of about 60 s (60-70% of PQH2).