The polypeptide composition of whole thylakoids and membrane subfragments was studied by using a modified two-dimensional gel electrophoresis technique of O'Farrell [J. Biol. Chem. 250,[4007][4008][4009][4010][4011][4012][4013][4014][4015][4016][4017][4018][4019][4020][4021] (1 975)]. The modifications were lithium dodecyl sulphate solubilization instead of SDS, reverse isofocusing and sensitive silver staining procedure. This high-resolution technique allowed us to separate and identify about 170 polypeptides of thylakoid membranes. After separating grana and stroma thylakoids it was found that both types of lamellae contained nearly equal amounts of polypeptides, but about 70 polypeptides were different in the two preparations. In grana thylakoids, 54 polypeptides out of 95 were found to be mainly present in grana and 31 of them were only present in grana preparations. In stroma membranes, 43 polypeptides out of 99 were mainly present in stroma lamellae and 38 of these polypeptides were exclusively present in stroma lamellae. In a functional photosystem I1 preparation, 61 individual polypeptides could be distinguished. Most of these polypeptides were present in both grana and stroma lamellae, but 22 of them were more pronounced in grana than in stroma lamellae. 9 polypeptides of photosystem I1 were distinctly different in grana and stroma lamellae, and these differences may connect closely with the functional differences of photosystem I1 in the two types of thylakoids.Our knowledge about the structural and functional organization of the photosynthetic membrane has been changed substantially during the last few years. Until the beginning of this decade it was the generally accepted concept that Photosystems (PS) I and 11 were located in stochiometrically equal amounts in the grana region of chloroplasts, while stroma exposed thylakoids contained mainly PS I only (1, 21. From the early '80s this concept has been dramatically changed. As an early step in this process it was recognized that the two photosystems do not contribute equally in the chloroplast lamellae of higher plants [3 -51.Other results have suggested that PS I is totally excluded from the grana partition region and is located exclusively in the stroma exposed thylakoids [6 -81. Recent evidence suggests that about 20% of PS I1 are still localized in stroma thylakoids [8,9], and these PS I1 are markedly different from those which are in the granal region [8, 91. The concept of PS I1 heterogeneity was developed to interpret the biphasic nature of the primary PS I1 photoactivity [lo-121. Different aspects of PS I1 heterogeneity have been addressed during the last few years and it is now believed that PS I1 occurs in the forms of two structural/functional configurations, termed PS 11, and PS 11, [S, 13, 141. Results obtained by the kinetic analysis of the primary photoactivity of fractionated thylakoids indicate that PS 11, is exclusively located in the membranes of grana partitions, while PS 11, and PS I are located in stroma exposed thylakoids [S,...