Missouri-Columbia, Columbia, Missouri 6521 1 (T.A.T.) localization of membrane proteins in the cyanobacterium Synechococcus sp. PCC7942 was determined by transmission electron microscopy utilizing immunocytochemistry with cells prepared by freeze-substitution. This preparation procedure maintained cellular morphology and permitted detection of cellular antigens with high sensitivity and low background. Synechococcus sp. PCC7942 is a unicellular cyanobacterium with thylakoids organized in concentric layers toward the periphery of the cell. Cytochrome oxidase was localized almost entirely in the cytoplasmic membrane, whereas a carotenoprotein (P35) was shown to be a cell wall component. The major photosystem I1 (PSII) proteins (Dl, D2, CP43, and CP47) were localized throughout the thylakoids. Proteins of the Cyt b6/f complex were found to have a similar distribution.Thylakoid luminal proteins, such as the Mn-stabilizing protein, were located primarily in the thylakoid, but a small, reproducible fraction was found in the outer compartment. The photosystem I (PSI) reaction center proteins and the ATP synthase proteins were found associated mostly with the outermost thylakoid and with the cytoplasmic membrane. These results indicated that the photosynthetic apparatus is not evenly distributed throughout the thylakoids. Rather, there is a radial asymmetry such that much of the PSI and the ATPase synthase is located in the outermost thylakoid. The relationship of this structure to the photosynthetic mechanism is discussed. It is suggested that the photosystems are separated because of kinetic differences between PSI1 and PSI, as hypothesized by H.-W. Trissl and c. Wilhelm (Trends Biochem Sci [1993] 18: 415-419).The structure of the photosynthetic membrane (the thylakoid) has been subjected to a wide variety of investigations (Albertsson et al., 1990;Olive and Vallon, 1991). The thylakoids in the chloroplasts of higher plants and algae are highly complex, specialized membranes consisting of two compartments, the grana and the stromal lamellae, which interconnect grana stacks. They perform interrelated photochemical and redox reactions, based on the functions of discrete supramolecular assemblies of proteins, pigments, and electron carriers. Distinctions are now made between appressed and
Growth of Synechococcus sp. strain PCC 7942 in iron-deficient media leads to the accumulation of an approximately 34-kDa protein. The gene encoding this protein, mapA (membrane-associated protein A), has been cloned and sequenced (GenBank accession number, L01621). The mapA transcript is not detectable in normally grown cultures but is stably accumulated by cells grown in iron-deficient media. However, the promoter sequence for this gene does not resemble other bacterial iron-regulated promoters described to date. The carboxyl-terminal region of the derived amino acid sequence of MapA resembles bacterial proteins involved in iron acquisition, whereas the amino-terminal end of MapA has a high degree of amino acid identity with the abundant, chloroplast envelope protein E37. An approach employing improved cellular fractionation techniques as well as electron microscopy and immunocytochemistry was essential in localizing MapA protein to the cytoplasmic membrane of Synechococcus sp. strain PCC 7942. When these cells were grown under iron-deficient conditions, a significant fraction of MapA could also be localized to the thylakoid membranes.
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