Oxysterol Induced Fluid-Fluid Phase Separation in Saturated Phosphatidylcholine Membranes: Influence of Lipid Chain Length.

We have recently reported the observation of closed-loop fluid-fluid immiscibility in 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) lipid membranes containing either 25-hydroxycholesterol (25HCH) or 27-hydroxycholesterol (27HCH) (Kamal et al. Proc Natl Acad Sci 120(25):2216002120, https://doi.org/10.1073/pnas.2216002120 ,  2023). Here we extend these studies to membranes of symmetric, saturated phospholipids having chain length varying from 12 to 16 carbon atoms. The phase behavior of the PC-25HCH and PC-27HCH membranes were probed by using fluorescence microscopy, atomic force microscopy and small angle X-ray scattering. 25HCH is found to induce fluid-fluid coexistence in these membranes for lipid chains lengths varying from 12 to 15 C atoms, whereas 27HCH induces similar phase behavior for chain lengths varying from 13 to 16. Thus the occurrence of fluid-fluid coexistence in these membranes depends both on the position of the -OH group in the oxysterol side chain as well as the membrane thickness. Our earlier studies have indicated that the phase behavior of these binary membranes can be understood in terms of the temperature-dependent orientation of the oxysterol in the membrane. Our present results suggest that the relative energies of these orientations depend on the membrane thickness and the oxysterol structure. Observation of fluid-fluid immiscibility in different saturated lipid model membranes containing 25HCH or 27HCH shows the generic nature of this phase behavior.