Abstract
The properties of bilayers composed of pure brain cerebroside (bCrb) or of binary mixtures of bCrb with brain ceramide, cholesterol, egg phosphatidylcholine or brain sphingomyelin have been studied using a combination of physical techniques. Pure bCrb exhibits a rather narrow gel-fluid transition centred at ≈65 °C, with a half-width at half-height T1/2 ≈ 3 °C. bCrb mixes well with both fluid and gel phospholipids and ceramide, and it rigidifies bilayers of egg phosphatidylcholine or brain sphingomyelin when the latter are in the fluid state. Cholesterol markedly widens the bCrb gel-fluid transition, while decreasing the associated transition enthalpy, in the manner of cholesterol mixtures with saturated phosphatidylcholines, or sphingomyelins. Laurdan and DPH fluorescence indicate the formation of fluid ordered phases in the bCrb:cholesterol mixtures. Macroscopic phase separation of more and less fluid domains is observed in giant unilamellar vesicles consisting of bCrb:egg phosphatidylcholine or bCrb:sphingomyelin. Crb capacity to induce bilayer permeabilization or transbilayer (flip-flop) lipid motion is much lower than those of ceramides. The mixtures explored here contained mostly bCrb concentrations >50 mol%, mimicking the situation of cell membranes in Gaucher’s disease, or of the Crb-enriched microdomains proposed to exist in healthy cell plasma membranes.
Highlights
Glycosphingolipids (GSL) are components of most eukaryotic cell plasma membranes
In the case of brain cerebroside (bCrb) the lower-melting component would arise from the lower-melting signal of non-hydroxylated cerebroside molecules, while the higher-melting component would be originated by the higher-melting non-hydroxylated signal plus the whole of the hydroxylated molecules
From the combined experiments described above, one can conclude that the main relevant properties of bCrb are (i) its capacity to rigidify fluid bilayers, (ii) its relatively good mixing with both fluid phospholipids and ceramides, and (iii) its interaction with Chol
Summary
Glycosphingolipids (GSL) are components of most eukaryotic cell plasma membranes. They consist of a ceramide backbone linked to a saccharide polar headgroup through an O-glycosidic linkage to the C1-hydroxyl of ceramide (Cer)[1] (Supplementary Figure S1). In most previous studies Crb were minority components in the various mixtures, while we have explored mixtures in which Crb is usually >50 mol%, corresponding in the ternary phase diagrams of Varela et al.[18] to the lower, right-hand region of the triangle. This may reflect the overall cell membrane situation in Gaucher’s disease, or the case of Crb-enriched microdomains expected to occur in healthy cell plasma membranes[3,4,5]. An additional novel aspect of our study is the comparative evaluation of SM, Crb and Cer as membrane permeabilizing agents and as inducers of trans-bilayer (flip-flop) motion, a number of observations with important physiological consequences in the cell
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