ISOLATION AND CHARACTERIZATION OF THYLAKOID VESICLES ENRICHED IN PEROXIDIZED LIPIDS

Thylakoid vesicles enriched in peroxidized lipids by 2- to 3-fold in c omparison with thylakoid membranes were isolated from the stroma of in tact chloroplasts obtained from primary leaves of two-week-old bean se edlings (Phaseolus vulgaris L. cv. Kinghorn). The fatty acid compositi on of the vesicles is similar to that of thylakoid membranes. The vesi cles also exhibit photosystem II activity, but the level of activity i s similar to 4-fold lower than that measured fcr thylakoid membranes. The chlorophyll a/b ratio for the vesicles is 6.59 +/- 0.21 compared w ith corresponding ratios of 2.85 +/- 0.02 and 9.85 +/- 0.62 for granal and stromal lamellae, respectively. It is clear from SDS-PAGE that th e polypeptide composition of the vesicles is distinguishable from that of thylakoid membranes, and immunological analyses of several thylako id membrane proteins have indicated that there are marked differences in protein composition between thylakoid vesicles and fractionated gra nal and stromal lamellae. Nonetheless, it is evident from their polype ptide and lipid composition that these vesicles originate from thylako ids, presumably by vesiculation. Moreover, the vesicles appear to be i n situ elements of the stroma inasmuch as they are isolated from intac t chloroplasts by a procedure that does not involve homogenization, an d similar vesicles are visible in situ by electron microscopy. Indeed, the finding that the vesicles are enriched in peroxidized lipids sugg ests that vesiculation may be a means of removing destabilizing lipid catabolites from thylakoid membranes.