Stoichiometry of pigments and radical pair formation under saturating pulse excitation in D1/D2/cytb(559) preparations

Chlorophyll a content and maximum yield of radical ion pair formation, P680(+.)Pheo(-.), were analyzed in D1/D2/cytb(559) preparations isolated from spinach. The following results were obtained: (a) chromatographic analysis with HPLC using an area-ratio method independent of extinction coefficients revealed that 7 +/- 0.5 Chl a are present per 2 Pheo; (b) based on highly sensitive silver staining of proteins and its densitometric evaluation, the contamination by CP47 was estimated to be 4%, i.e., no more than 0.8 Chl can be ascribed to this Chl a binding protein; (c) measurements of laser flash induced absorption changes at 830 nm as a function of the pulse energy indicate that under light saturating conditions one P680(+.)Pheo(-.) radical pair can be formed per 7-8 Chl a; (d) based on the saturation behavior of the different decay components of the absorption changes and on a comparison of the absorption and single photon counting fluorescence data, the 5-ns component is inferred to originate from 'disconnected' Chi. These results lead to the conclusion that functionally fully competent D1/D2/cytb(559) complexes contain 6 Chl per 2 Pheo. More than 90% of the centres are able to perform a charge separation leading to the radical pair P680(+.)Pheo(-.).