Summary
Homogeneous, small, single-bilayer vesicles were prepared from egg phosphatidylcholine with various concentrations of cholesterol by ultrasonic dispersion in 0.1m KCl, 0.01m Tris, pH 8.0, buffer, followed by gel chromatography. The shape and size distributions of the fractionated vesicles were investigated for preparations with cholesterol compositions from 0 to 50 moles/100 moles, using freeze-etch electron microscopy. The size distribution was estimated from the shadow width of vesicles which were exposed by etching and the vesicle shape was checked by comparing the images obtained by tilting the replicas. The widths of the vesicle diameter distributions were relatively broad, corresponding to standard deviations in the range 60–90 Å, but showing no systematic variation with cholesterol composition. In all cases it was found that 70% of the vesicle diameters lay within 150 Å of the modal value. The apparent vesicle diameters remained constant for cholesterol compositions up to 20 moles/100 moles (modal diameter=330 ± 20 Å, mean diameter = 350 ± 3 Å), but there was a sharp net increase in diameter at 30 moles cholesterol/100 moles reaching a model diameter of 430 ± 20 Å (mean diameter = 430 ± 3 Å) at 50 moles cholesterol/100 moles. Using the tilted microscope stage it was found that all vesicles were spherical at all cholesterol compositions studied, including those above 30 moles cholesterol/100 moles. The molecular mechanism by which cholesterol controls the vesicle size is discussed in terms of the asymmetric distribution of cholesterol across the vesicle bilayer.
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References
Bernard, L. de. 1958. Associations moléculaires entre les lipides. II.—Lécithine et cholésterol.Bull. Soc. Chim. Biol. 40:161
Chrzeszczyk, A., Wishnia, A., Springer, C.S. 1977. The intrinsic structural asymmetry of highly curved phospholipid bilayer membranes.Biochim. Biophys. Acta 470:161
Demel, R.A., Geurts van Kessel, W.S.M., Deenen, L.L.M. van 1972. The properties of polyunsaturated lecithins in monolayers and liposomes and the interactions of these lecithins with cholesterol.Biochim. Biophys. Acta 266:24
Freeman, R., Finean, J.B. 1975. Cholesterol-lecithin association at molar ratios of up to 2∶1.Chem. Phys. Lipids 14:313
Gent, M.P.N., Prestegard, J.H. 1974. Cholesterol-phosphatidylcholine interactions in vesicle systems. Implication of vesicle size and proton magnetic resonance linewidth changes.Biochemistry 13:4027
Huang, C. 1969. Studies on phosphatidylcholine vesicles. Formation and physical characteristics.Biochemistry 8:344
Huang, C., Charlton, J.P. 1972. Studies on the state of phosphatidylcholine molecules before and after ultrasonic and gel-filtration treatments.Biochem. Biophys. Res. Commun. 46:1660
Huang, C., Mason, J.T. 1978. Geometric packing constraints in egg phosphatidylcholine vesicles.Proc. Nat. Acad. Sci. USA 75:308
Huang, C., Sipe, J.P., Chow, S.T., Martin, R.B. 1974. Differential interaction of cholesterol with phosphatidylcholine on the inner and outer surfaces of lipid bilayer vesicles.Proc. Nat. Acad. Sci. USA 71:359
Johnson, S.M. 1973. The effect of charge and cholesterol on the size and thickness of sonicated phospholipid vesicles.Biochim. Biophys. Acta 307:27
Kruijff, B. de, Cullis, P.R., Radda, G.K. 1975. Differential scanning calorimetry and31P NMR studies on sonicated and unsonicated phosphatidylcholine liposomes.Biochim. Biophys. Acta 406:6
Kruijff, B. de, Cullis, P.R., Radda, G.K. 1976. Outside-inside distributions and sizes of mixed phosphatidylcholine-cholesterol vesicles.Biochim. Biophys. Acta 436:729
Ladbrooke, B.D., Williams, R.M., Chapman, D. 1968. Studies on lecithin-cholesterol-water interactions by differential scanning calorimetry and x-ray diffraction.Biochim. Biophys. Acta 150:333
Lecuyer, H., Dervichian, D.G. 1969. Structure of aqueous mixtures of lecithin and cholesterol,J. Mol. Biol. 45:39
Levine, Y.K., Wilkins, M.H.F. 1971. Structure of oriented lipid bilayers.Nature New Biol. 230:69
Marsh, D. 1974. An interacting spin label study of lateral expansion in dipalmitoyllecithin-cholesterol bilayers.Biochim. Biophys. Acta 363:373
Marsh, D., Phillips, A.D., Watts, A., Knowles, P.F. 1972. A spin label study of fractionated egg phosphatidylcholine vesicles.Biochem. Biophys. Res. Commun. 49:641
Marsh, D., Smith, I.C.P. 1973. An interacting spin label study of the fluidizing and condensing effects of cholesterol on lecithin bilayers.Biochim. Biophys. Acta 298:133
Newman, G.C., Huang, C. 1975. Structural studies on phosphatidylcholine-cholesterol mixed vesicles.Biochemistry 14:3363
Schreier-Muccillo, S., Marsh, D., Dugas, H., Schneider, H., Smith, I.C.P. 1973. A spin probe study of the influence of cholesterol on motion and orientation of phospholipids in oriented multibilayers and vesicles.Chem. Phys. Lipids 10:11
Shah, D.O., Schulman, J.H. 1967. Influence of calcium, cholesterol and unsaturation on lecithin monolayers.J. Lipid Res. 8:215
Shimshick, E.J., McConnell, H.M. 1973. Lateral phase separations in binary mixtures of cholesterol and phospholipids.Biochem. Biophys. Res. Commun. 53:446
Watts, A., Marsh, D., Knowles, P.F. 1978. Characterization of dimyristoyl phosphatidylcholine vesicles and their dimensional changes through the phase transition. Molecular control of membrane morphology.Biochemistry (in press)
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Forge, A., Knowles, P.F. & Marsh, D. Morphology of egg phosphatidylcholine-cholesterol single-bilayer vesicles, studied by freeze-etch electron microscopy. J. Membrain Biol. 41, 249–263 (1978). https://doi.org/10.1007/BF01870432
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DOI: https://doi.org/10.1007/BF01870432