Biophys J, February 2002, p. 813-822, Vol. 82, No. 2

Cyclodextrin-Induced Lipid Lateral Separation in DMPC Membranes: ²H Nuclear Magnetic Resonance Study

Michel Roux,^* Rachel Auzely-Velty, Florence Djedaini-Pilard, and Bruno Perly

 ^*Section de Biophysique des Protéines et des Membranes, Département de Biologie Cellulaire et Moléculaire URA CNRS 2096,  Service de Chimie Moléculaire, Département de Recherche sur l'État Condensé, les Atomes et les Molécules and CEA Saclay, 91191 Gif sur Yvette cedex, France

Cholesteryl cyclodextrins, obtained by grafting a cholesterol moiety on the oligosaccharide core, combine the size selectivity of the cyclodextrin cavity with the carrier properties of model membrane systems such as micelles or liposomes. The cholesteryl cyclodextrins were incorporated as guests in chain perdeuterated dimyristoyl phosphatidylcholine (DMPC-d54) membranes. The deuterium nuclear magnetic resonance (NMR) spectra obtained with the A form of cholesteryl--cyclodextrin (CC_A), with a succinyl spacer inserted between the cholesterol moiety and the cyclodextrin headgroup, indicated that this compound induces a lateral phase separation of DMPC-d54, into a pure lipid phase and a cholesteryl cyclodextrin-rich phase. The lipid exchange rate between the two phases was slow on the NMR timescale (>10⁵ s), and two well-resolved spectral components could be detected. The laterally segregated mixed phase was observed at various membrane concentrations of cholesteryl cyclodextrin, even with dispersions containing only 5% of the derivative. The dePaked spectra allowed the determination of the relative amount of DMPC-d54 molecules contained in each phase, giving ~1 to 1.5 DMPC molecules per unit of CC_A. This ratio was found to be independent of the total membrane concentration of CC_A. The cholesteryl cylodextrin-rich phase was detected on a large range of temperature from 12°C to 25°C and exhibits a smooth transition from a fluid environment to a more ordered state, occurring ~0°C. A boundary phase between the pure lipid and cyclodextrin-rich phase was detected at 19°C just below the fluid-to-gel transition. The average orientational order was reduced in the cholesteryl cyclodextrin-rich phase, and quasi-independent of temperature, as opposed to the order parameters measured for the NMR signals of the pure lipid phase. However, the NMR data obtained with CC_A deuterated on the cyclodextrin headgroup indicated that the latter was quasistatic, with very large order parameters (~120 kHz) at all temperatures, suggesting strong interactions between neighboring cyclodextrin headgroups. The interactions of DMPC-d54 membranes with the B form of cholesteryl--cyclodextrin, lacking the succinyl spacer, was also investigated in a parallel study. No lateral phase separation was found with this compound, indicating that the spatial location and a precise positioning (allowed by the spacer) of the cyclodextrin headgroup at the membrane interface was crucial for the stability of the cholesteryl cyclodextrin lamellar phase.

Biophys J, February 2002, p. 813-822, Vol. 82, No. 2
© 2002 by the Biophysical Society   0006-3495/02/02/813/10  $2.00

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