Lipid Lateral Segregation driven by Diacyl Cyclodextrin Interactions at the Membrane Surface

¹ CNRS URA 2096
² OZ-Biosciences
³ Centre d'Etudes de Saclay
⁴ Université de Picardie Jules Verne

^* To whom correspondence should be addressed. E-mail: michel.roux{at}cea.fr.

Submitted on November 22, 2006
Revised on January 11, 2007
Accepted on 30 April 2007

	Abstract

Cyclodextrins are hydrophilic molecular cages with an hydrophobic interior allowing the inclusion of water-insoluble drugs. Amphiphilic cyclodextrins obtained by appending an hydrophobic anchor, were designed to improve the cell targeting of the drug-containing cavities through their liposome transportation in the organism. After insertion in model membranes, they were found to induce a lateral phase separation into a pure lipid phase and a fluid cyclodextrin-rich phase (L_CD) with reduced acyl chain order parameters, as observed with a derivative containing a cholesterol anchor (M. Roux, R. Auzely-Velty, F. Djedaïni-Pilard, & B. Perly. 2002. Biophysical Journal, 8:813-822). We present another class of amphiphilic cyclodextrins obtained by grafting aspartic acid esterified by two lauryl chains on the oligosaccharide core via a succinyl spacer. The obtained dilauryl--cyclodextrin (DLC) was inserted in chain perdeuterated dimyristoyl phosphatidylcholine (DMPC-d54) membranes and studied by deuterium nuclear magnetic resonance (²H-NMR). A laterally-segregated mixed phase was found to sequester three times more lipids than the cholesteryl derivative (~ 4-5 lipid per monomer of DLC) and a quasi pure L_CD phase could be obtained with a 20% molar concentration of dilauryl--cyclodextrin. When cooled below the main fluid-to-gel transition of DMPC-d54 the DLC-rich phase stays fluid, coexisting with pure lipid in the gel state, and exhibits a sharp transition to a gel phase with frozen DMPC acyl chains at 12.5°C. No lateral phase separation was observed with partially or fully methylated dilauryl--cyclodextrin, confirming that the stability of the segregated L_CD phase was governed through hydrogen bonds-mediated intermolecular interactions between cyclodextrin headgroups at the membrane surface. As opposed to native dilauryl--cyclodextrin, the methylated derivatives were found to strongly increase the orientational order of DMPC acyl chains as the temperature reaches the membrane fluid-to-gel transition. The results are discussed in relation to the "anomalous swelling" of saturated phosphatidylcholine multilamellar membranes known to occur in the vicinity of the main fluid-to-gel transition.

Key Words: 2H-NMR, DMPC, cyclodextrin, lateral segregation, membrane, microdomain