Photochemical activity and structural studies of photosystems derived from chloroplast grana and stroma lamellae

Stroma lamellae isolated by French press treatment, and Photosystem I and II fractions isolated by digitonin treatment of the grana fraction (free of stroma lamellae), were compared with respect to electron transport activities and ultrastructural characteristics. The results show:. 1. About 15 % of the chlorophyll in mature spinach leaf chloroplasts occurred in the stroma lamellae. The separated grana membranes, when fractionated with digitonin, were comprised of 60 % Photosystem II and 40 % Photosystem I (on a chlorophyll basis), with essentially complete recovery of the starting material. 2. Action spectra and electron transport data for characteristic Photosystem I and II activities supported the concept that NADP⁺ reduction activity was closely correlated with the content of P700 in the fractions. In no instance were there significant rates of NADP⁺ reduction from water or an alternative Photosystem II donor (diphenylcarbazide) in the Photosystem II enriched fraction, beyond that which could be entirely accounted for by the residual P700 content. 3. The grana Photosystem II fraction, low in P700 and water or diphenyl carbazide→ NADP⁺ electron transport activity, could be recombined with the grana Photosystem I fraction and the reconstituted preparations were then capable of increased rates of diphenyl carbazide→ NADP⁺ reduction which were sensitive to a Photosystem II inhibitor, dichlorophenylmethylurea (DCMU). Stroma lamellae Photosystem I fraction, prepared by digitonin treatment was not capable of recombining with the grana Photosystem II fraction. For the grana, the results are most simply explained by the series electron transport scheme which assumes that NADP⁺ reduction from water normally proceeds via a series interaction between Photosystem II and a P700-containing Photosystem I. However, it cannot be unequivocally stated that the electrons from Photosystem II pass through P700 since none of our measurements followed P700 turnover directly. 4. Photosystem I obtained from stroma and grana membranes were quite similar with regard to electron transport activity, P700 content, ultrastructure appearance and ultrafiltration characteristics, and differing in that stroma Photosystem I fragments did not recombine with the grana Photosystem II fraction and reconstitute electrons transport activity from diphenyl carbazide→ NADP⁺. Plastocyanin was present in both grana and stroma membranes as shown by a sensitive bioassay. 5. The data are consistent with the chloroplast membrane model proposed earlier by Arntzen, Dilley and Crane [J. Cell. Biol., 43 (1969) 16] as regards the binary nature of the grana membranes, with Photosystem I localized on the outer "half" and Photosystem II on the inner "half" of a 90-Å grana thylakoid membrane.