Cytochrome f/b6 and ATP synthetase (CF0-CF1) complexes from spinach chloroplasts have been reconstituted into liposomes prepared from soybean phospholipids and purified spinach galactolipids. Freeze-fracture analysis revealed homogeneous populations of particles spanning the lipid bilayers with their elongated axes perpendicular to the membrane plane. The lipid composition of the liposomes had no effect on the size of the reconstituted complexes, the average diameter of cytochrome f/b6 complex measuring 8.5 nm, and of the CF0 base piece of the ATP synthetase, 9.5 nm. When reconstituted cytochrome f/b6 complexes were cross-linked by means of antibodies prepared against the whole complex, the thus aggregated particles formed either hexagonal or square arrays. In both instances the center-to-center spacing of the particles was 8.3 nm, thereby suggesting that this value could be closer to the real diameter of the complexes than the one obtained from measuring individual particles. Assuming an ellipsoidal shape for these particles, and using a measured height of 11 nm, a molecular weight of approximately 280,000 could be calculated for the reconstituted cytochrome f/b6 complex, consistent with a dimeric configuration. In many instances the crystalline sheets of antibody-aggregated cytochrome f/b6 complexes were found to be free in the buffer solution; apparently the antibody-induced strains caused the sheet-like aggregates to pop out of the liposomal membranes. Agglutination studies of inside-out and right-side-out thylakoid vesicles revealed the antigenic determinants of the cytochrome f and cytochrome b6 polypeptides to be exposed on the inner thylakoid surface and to be present in stacked and unstacked membrane regions. The molecular weight calculated from the size of freeze-fractured CF0 base pieces was over twice the value determined by x-ray scattering data. This discrepancy may be caused by significant lipid domains within the base piece, or by an unusual fracturing behavior of the base piece in reconstituted liposomes.
Reconstitution of cytochrome f/b6 and CF0-CF1 ATP synthetase complexes into phospholipid and galactolipid liposomes.
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E Mörschel, L A Staehelin; Reconstitution of cytochrome f/b6 and CF0-CF1 ATP synthetase complexes into phospholipid and galactolipid liposomes.. J Cell Biol 1 August 1983; 97 (2): 301–310. doi: https://doi.org/10.1083/jcb.97.2.301
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