Oxygen-evolving diatom thylakoid membranes

AbstractTwo protocols were developed that yielded purified oxygen-evolving thylakoid membranes from the diatom Cylindrotheca fusiformis. One protocol employed sonication, while the second involved French press lysis of protoplasts formed by brief culture of cells in a cation-depleted medium. Regardless of the method of cell breakage, some damage to electron transport components occurred. For preservation of both light-dependent electron transport activity and in vivo fluorescence properties, 2 M sorbitol proved to be more effective than 1 M sorbitol, regardless of the method used for cell lysis. Thylakoids purified in 2 M sorbitol using the protoplast/French press method showed the best preservation of in vivo fluorescence emission signals and Photosystem II activity with ferricyanide was completely inhibited by DCMU. Thylakoids purified in 2 M sorbitol using sonication had higher rates of Photosystem II activity with ferricyanide, but this activity was less sensitive to DCMU. Whole-chain electron transport activity was low in all preparations. This is the first report of O2 evolution and of long-wavelength fluorescence in purified thylakoids of any chromophytic alga