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Fluid-Phase Chain Unsaturation Controlling Domain Microstructure and Phase in Ternary Lipid Bilayers Containing GalCer and Cholesterol

Wan-Chen Lin ^*, Craig D. Blanchette ^*  and Marjorie L. Longo 

^* Biophysics Graduate Group, University of California, Davis, California; Biophysical and Interfacial Science Group, Chemistry and Material Science, Lawrence Livermore National Laboratory, Livermore, California; and Department of Chemical Engineering and Materials Science, University of California, Davis, California

Correspondence: Address reprint requests to Marjorie L. Longo, Dept. of Chemical Engineering and Materials Science, University of California, Davis, CA 95616. Tel.: 530-754-6348; Fax: 530-752-1031; E-mail: mllongo{at}ucdavis.edu.

We report the microstructure and phase behavior of three ternary mixtures each containing a long-chain saturated glycosphingolipid, galactosylceramide (GalCer), and cholesterol at room temperature. The unsaturation level of the fluid-phase component was varied by lipid choice, i.e., saturated 1,2-dilauroyl-sn-glycero-3-phosphocholine (DLPC), singly unsaturated 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), or doubly unsaturated 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). GalCer was used because of its biological significance, for example, as a ligand in the sexual transmission of HIV and stimulator of natural killer T-cells. Supported lipid bilayers of the ternary mixtures were imaged by atomic force microscopy and GalCer-rich domains were characterized by area/perimeter ratios (A/P). GalCer domain phase transitions from solid (S) to liquid (L) phase were verified by domain behavior in giant unilamellar vesicles, which displayed two-dimensional microstructure similar to that of supported lipid bilayers. As cholesterol concentration was increased, we observed 2.5, 10, and 20-fold decreases in GalCer domain A/P for bilayers in L-S phase coexistence containing DOPC, POPC, and DLPC, respectively. The transition to L-L phase coexistence occurred at 10 mol % cholesterol for bilayers containing DOPC or POPC and was accompanied by maintenance of a constant A/P. L-L phase coexistence did not occur for bilayers containing DLPC. We systematically relate our results to the impact of chain unsaturation on the interaction of the fluid-phase lipid and cholesterol. Physiologically, these observations may give insight into the interplay of fatty acid chain unsaturation, sterol concentration, and lipid hydrophobic mismatch in membrane phenomena.

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