ISOLATION OF PHOTOSYSTEM-II ENRICHED MEMBRANE-VESICLES FROM SPINACH-CHLOROPLASTS BY PHASE PARTITION

Partition in an aqueous Dextran-polyethylene glycol 2-phase systems was used for the separation of chloroplast membrane vesicles obtained by press treatment of a grana-enriched fraction after unstacking in a low salt buffer. The fractions obtained were analyzed with respect to chlorophyll, photochemical activities and ultrastructural characteristics. The material partitioning to the Dextran-rich bottom phase consisted of large membrane vesicles possessing mainly Photosystem II properties with low contribution from Photosystem I. Measurements of the H2O to phenyl-p-benzoquinone and ascorbate-Cl2Ind [2,6-dichlorophenolindophenol] to NADP+ electron transport rates indicate a ratio of around 6 between Photosystem II and I. The total fractionation procedure was completed within 2-3 h with high recovery of both the Photosystem II water-splitting activity and the Photosystem I reduction of NADP+. These data demonstrate that press treatment of low-salt destabilized grana membranes yields a population of highly Photosystem-II enriched membrane vesicles which can be discriminated by the phase system. Such membrane vesicles probably originate from large regions in the native grana membrane which contain virtually only Photosystem II.