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We employed small-angle X-ray scattering measurement using synchrotron radiation to study structural properties of monosialoganglioside (GM1)/cholesterol (chol) mixtures in aqueous dispersions, in the presence of 0-50 mM CaCl2. Various molar ratios of GM1/chol were examined from 1:0 to 1:5. Without CaCl2 the radius of gyration (Rg) of the aggregate increased linearly with increasing cholesterol content up to GM1/chol = 1/1. At cholesterol content above GM1/chol = 1/1, Rg of the aggregate was almost constant, and a sharp diffraction peak at q = 0.18 Å-1, characteristic of cholesterol monohydrate crystals, was observed, suggesting the solubility limit of cholesterol in GM1 aggregates is about GM1/chol = 1/1. For the samples containing cholesterol of GM1/chol = 1/0.6 and above, the diffraction patterns showed a set of peaks that correspond to ~85 Å periodic lamellar structures in the presence of CaCl2 at concentrations higher than ~10 mM. The observed lamellar peak positions and the intensity ratio of the lamellar peaks (1st-4th) was not changed by the addition of CaCl2 up to 100 mM, demonstrating once formed multilayered structure of GM1/chol was not altered under the present conditions. Electron density profiles for this GM1/chol bilayer were obtained. The Ca2+-induced vesicle-to-lamellar phase separation of this system was found to be highly reversible by the Ca2+ chelation with ethylenediamine-tetraacetic acid.

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