Abstract
There is a significant amount of experimental data, obtained predominantly from fluorescence studies, showing that sterol-containing liposomes can exhibit multiple biphasic changes in membrane properties at specific critical mole fractions of sterol such as 20.0, 22.2, 25.0, 33.3, 40.0, and 50.0 mol%. This can be understood in terms of the sterol regular distribution (e.g., superlattice) model. Here, the authors use excitation generalized polarization of 6-lauroyl-2-dimethylamino-naphthalene fluorescence in fluid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine/cholesterol unilamellar vesicles to illustrate the experimental procedures and conditions that are required to detect multiple biphasic changes at predicted sterol mole fraction values in liposomal membranes. For this detection, the use of small sterol increments over a wide sterol mole fraction range is essential. Lipid concentration, incubation time, thermal history, and degree of sterol oxidation of liposomal membranes are critical factors. The principles and methodologies described here can be extended to other probes or bioactive molecules, such as enzymes, and can be applied to study sterol lateral organization in multicomponent lipid membranes.
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References
Simons, K. and Ikonen, E. (1997) Functional rafts in cell membranes. Nature 387, 569–572.
Chong, P. L.-G. and Sugar, I. P. (2002) Fluorescence studies of lipid regular distribution in membranes. Chem. Phys. Lipids 116, 153–175.
Chong, P. L.-G. and Olsher, M. (2004) Fluorescence studies of the existence and functional importance of regular distributions in liposomal membranes. Soft Mater. 2, 85–108.
Somerharju, P., Virtanen, J. A., and Cheng, K. H. (1999) Lateral organisation of membrane lipids: the superlattice view. Biochim. Biophys. Acta 1440, 32–48.
Chong, P. L.-G. (1994) Evidence for regular distribution of sterols in liquid crystalline phosphatidylcholine bilayers. Proc. Natl. Acad. Sci., U.S.A. 91, 10,069–10,073.
Virtanen, J. A., Ruonala, M., Vauhkonen, M., and Somerharju, P. (1995) Lateral organization of liquid-crystalline cholesterol-dimyristoylphosphatidylcholine bilayers. Evidence for domains with hexagonal and centered rectangular cholesterol superlattices. Biochemistry 34, 11,568–11,581.
Chong, P. L.-G., Liu, F., Wang, M. M., Truong, K., Sugar, I. P., and Brown, R. E. (1996) Fluorescence evidence for cholesterol regular distribution in phosphatidylcholine and in sphingomyelin lipid bilayers. J. Fluoresc. 6, 221–230.
Liu, F., Sugar, I. P., and Chong, P. L.-G. (1997) Cholesterol and ergosterol superlattices in three-component liquid crystalline lipid bilayers as revealed by dehydroergosterol fluorescence. Biophys. J. 72, 2243–2254.
Wang, M. M., Sugar, I. P., and Chong, P. L.-G. (1998) Role of the sterol superlattice in the partitioning of the antifungal drug nystatin into lipid membranes. Biochemistry 37, 11,797–11,805.
Liu, F. and Chong, P. L.-G. (1999) Evidence for a regulatory role of cholesterol superlattices in the hydrolytic activity of secretory phospholipase A2 in lipid membranes. Biochemistry 38, 3867–3873.
Wang, M. M., Sugar, I. P., and Chong, P. L.-G. (2002) Effect of double bond position on dehydroergosterol fluorescence intensity dips in phosphatidylcholine bilayers with saturated sn-1 and monoenoic sn-2 acyl chains. J. Phys. Chem. 106, 6338–6345.
Wang, M. M., Olsher, M., Sugar, I. P., and Chong, P. L.-G. (2004) Cholesterol superlattice modulates the activity of cholesterol oxidase in lipid membranes. Biochemistry 43, 2159–2166.
Parker, A., Miles, K., Cheng, K. H., and Huang, J. (2004) Lateral distribution of cholesterol in dioleoylphosphatidylcholine lipid bilayers: cholesterol-phospholipid interactions at high cholesterol limit. Biophys. J. 86, 1532–1544.
Olsher, M., Yoon, S. I., and Chong, P. L.-G. (2005) Role of sterol superlattice in free radical-induced sterol oxidation in lipid membranes. Biochemistry 44, 2080–2087.
Chong, P. L.-G. (1996) Membrane-free volume variation with bulky lipid concentration by regular distribution: a functionally important property explored by pressure studies of phosphatidylcholine bilayers in High-Pressure Effects in Molecular Biophysics and Enzymology, (Markley, J. L., Northrop, D. B., and Royer, C. A., eds.), Oxford University Press, New York, NY, pp. 298–313.
Huang, J. and Feigenson, G. W. (1999) A microscopic interaction model of maximum solubility of cholesterol in lipid bilayers. Biophys. J. 76, 2142–2157.
Parasassi, T. G. and Gratton, E. (1995) Membrane lipid domains and dynamics as detected by laurdan fluorescence. J. Fluoresc. 5, 59–69.
Cannon, B., Heath, G., Huang, J., Somerharju, P., Virtanen, J. A., and Cheng, K. H. (2003) Timeresolved fluorescence and Fourier transform infrared spectroscopic investigations of lateral packing defects and superlattice domains in compositionally uniform phosphatidylcholine bilayers. Biophys. J. 84, 3777–3791.
Bartlett, G. R. (1959) Phosphorus assay in column chromatography. J. Biol. Chem. 234, 466–468.
Chong, P. L.-G. and Wong, P. T. T. (1993) Interactions of laurdan with phosphatidylcholine liposomes: a high pressure FTIR study. Biochim. Biophys. Acta 1149, 260–266.
Chong, P. L.-G. and Thompson, T. E. (1986) Depolarization of dehydroergosterol in phospholipid bilayers. Biochim. Biophys. Acta 863, 53–62.
Lin, H. and Huang, C. (1988) Eutectic phase behavior of 1-stearoyl-2-caprylphosphatidylcholine and dimyristoylphosphatidylcholine mixtures. Biochim. Biophys. Acta 946, 178–184.
Chong, P. L.-G., Tang, D., and Sugar, I. P. (1994) Exploration of physical principles underlying lipid regular distribution: effects of pressure, temperature, and radius of curvature on E/M dips in pyrene-labeled PC/DMPC binary mixtures. Biophys. J. 66, 2029–2038.
Venegas, B., Sugar, I., P., and Chong, P. L.-G. (2007) Critical factors for detection of biphasic changes in membrane properties at specific sterol mole fractions for maximal superlattice formation. J. Phys. Chem. B. 111, 5180–5192.
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Chong, P.LG., Venegas, B., Olsher, M. (2007). Fluorescence Detection of Signs of Sterol Superlattice Formation in Lipid Membranes. In: Dopico, A.M. (eds) Methods in Membrane Lipids. Methods in Molecular Biology™, vol 400. Humana Press. https://doi.org/10.1007/978-1-59745-519-0_11
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DOI: https://doi.org/10.1007/978-1-59745-519-0_11
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