ISOLATION AND CHARACTERIZATION OF THE PHOTOSYSTEM 2 REACTION-CENTER COMPLEX FROM A DOUBLE MUTANT OF CHLAMYDOMONAS-REINHARDTII

A rapid procedure has been developed for the isolation of the photosys tem two reaction centre complex (PS II RC) from a double mutant of Chl amydomonas reinhardtii, F54-14, which lacks the Photosystem one comple x and the chloroplast ATPase. Thylakoid membranes are solubilised with 1.5% (w/v) Triton X-100 and the PS II RC purified by anion-exchange c hromatography using TSK DEAE-650(S) (Merck). The complex has a pigment stoichiometry of approximately six chlorophyll a: two pheophytin a: o ne cytochrome b-559: one to two beta-carotene. It photoaccumulates red uced pheophytin and oxidised P680 in the presence of sodium dithionite and silicomolybdate, respectively. Immunoblotting experiments have co nfirmed the presence of the D1 and D2 polypeptides in this complex. Th e cr-subunit of cytochrome b-559 was identified by N-terminal sequenci ng. Comparison of the complex with the PS II RC from pea using SDS-pol yacrylamide gel electrophoresis showed that their polypeptide composit ions were similar. However, the alpha-subunit of cytochrome b-559 from C. reinhardtii has a lower apparent molecular weight than the pea cou nterpart whereas the beta-subunit is larger.