Vesicles prepared with the French press from membranes of cyanelles of Cyanophora paradoxa retain 02 evolution activity with rates up to 500 micromoles 2,6-dichlorophenolindophenol reduced per hour per milligram chlorophyll. This activity is immediately lost when the vesicles are transferred from the sucrose-phosphate-citrate preparation buffer into dilute phosphate buffer. Similar preparations from Phormidium laminosum, a thermophilic cyanobacterium retain activity under such conditions. Photosystem I activities of both cyanobacterial vesicle preparations were determined by direct spectrophotometric measurement of N,N,N',N'-tetramethyl-p-phenylenediamine photooxidation in the presence of anthraquinone-2-sulfonate. The rates so determined were compared with rates of 02 taken up in the presence of methyl viologen or anthraquinone-2-sulfonate as electron acceptors. The predicted stoichiometry of two was observed for moles of N,N,N',N'-tetramethyl-p-phenylenediamine TMPD is readily reduced by PSII and oxidized by PSI, as shown by the fact that it overcomes DBMIB inhibition of NADP photoreduction coupled to water oxidation (12). In this case, it serves as a bypass for electron flow past the DBMIB inhibition site. In spinach chloroplasts, TMPD reacts with plastocyanin, which serves as a direct donor to PSI (3). In this study we report on the PSII activities of cyanobacterial preparations as well as the direct spectrophotometric measurement of TMPD photooxidation by PSI. The rates observed for PSI in this manner are compared to the rates measured in the traditional way as 02 uptake with excess ascorbate present. The impetus for the study was our interest in the photoreactions of the cyanelle of Cyanophora paradoxa, which although an endosymbiotic organelle, has the photochemical capabilities of cyanobacteria. The lability of the cyanelle PSII is shown as well as the ability of PSI to use ascorbate itself as a donor to a significant extent. This is in contrast to other photosynthetic systems in which ascorbate is a poor donor to PSI (4).
MATERIALS AND METHODSSpinach was purchased from a local market. The methods used for growth of C. paradoxa and for isolation of the cyanelle were those previously described (8). The original culture of Phormidium laminosum (OH1-P), a gift from Dr. Richard Castenholz, was grown as described by Jackson and Castenholz (5). Spinach chloroplasts were prepared as previously reported (6) and finally suspended in 0.4 M sorbital and 0.1 M Tris buffer (pH 7.0). Photosynthetically active vesicles were prepared from cyanelles and from P. laminosum cells by treatment in a French Press at 14,000 p.s.i. in the medium described by Dilworth and Gantt (12) which contained 0.5 M sucrose, 0.5 M K-phosphate and 0.3 M sodium citrate, all at pH 7.0. This is designated SPC buffer. Serum albumin was not added, as was previously done by Katoh and Gantt (7). Better activity was observed for PSII if the final vesicle preparation contained 0.2 mg Chl/ml or more.PSII was assayed by DPIP photoreduction in...