How do Cholesterol and Saturated Sphingolipids Affect Acyl Chain Order in the Fluid Phase of Binary POPC Bilayers - a Study with 1-oleoyl-2-propionyl-DPH-sn-glycero-3-phosphocholine

Abstract

It is known that cholesterol increases the order in the fluid phase of a lipid bilayer but it has remained unclear whether or not various sphingolipids also have an ordering effect on the bulk phase ordering properties in the fluid phase. The acyl chain order in the gel phases has previously been reported with trans-parinic acid (tPA; steady state anisotropy or lifetime analysis) but acyl chain order in the fluid phase phase separated systems has been mostly measured indirectly, e.g. with 1,6-Diphenyl-1,-3,5-hexatriene (DPH) or N-rhodamine.dipalmitoylphosphatidyletanolamine (Rho-DOPE). By using a 1-oleoyl-2-propionyl-DPH-sn-glycero-3-phosphocholine (18:1-DPH-PC) as a reporter molecule the order in the fluid phase could be measured since the unsaturated probe has preferential partitioning into the disordered phase. The systems we studied were: POPC with 50 mol% palmitoyl sphingomyelin (PSM), POPC with 29 mol% of either palmitoyl ceramide (PCer), palmitoyl galactosyl ceramide (PGalCer) or palmitoyl glucosyl ceramide (PGluCer). In the above bilayer systems (with coexisting disordered and ordered phases present), 18:1-DPH-PC steady state anisotropy reported no gel phase melting during temperature ramps. This contrasts with tPA, whose anisotropy in binary bilayers reported meltings of the ordered phase. 18:1-DPH-PC steady state anisotropy measurements at 23°C showed that PCer, PGlcCer, PGalCer addition to a fluid POPC bilayer had a very minor ordering effect compared to cholesterol addition. These results indicate that cholesterol is superior to saturated sphingolipids to increase order in the fluid phase and that 18:1-DPH-PC is a fluorophore suitable to measure membrane order in the fluid phase.