PHASE COEXISTENCE IN MODEL LIPID MEMBRANES MEASURED BY LAURDAN FLUORESCENCE

Abstract

The phase state of the lipid bilayer greatly influences membrane function and membrane-related protein activity. The spectral sensitivity of 2-dimethylamino-6-lauroylnaphthalene (Laurdan) to the polarity of its immediate environment was used to study phase coexistence in binary and ternary model membrane systems at various temperatures and cholesterol concentrations. Due to increased dipolar relaxation in the liquid-disordered phase, Laurdan displays a 50-nm bathochromic spectral shift compared to spectra indicative of the liquid-ordered phase. We have found that low temperatures can have an ordering spectral bias on samples that are in the pure liquid-disordered phase, therefore partitioning into pure liquid-ordered and liquid-disordered spectra is insufficient in quantifying regions of phase coexistence in binary systems.

Consequently, we have detailed a method for using temperature correspondences between samples of palmitoyl-oleyl phosphatidylcholine (POPC) / cholesterol and brain sphingomyelin (BSM) / cholesterol with samples of dimyristoyl-phosphatidylcholine (DMPC) / cholesterol to map phase boundaries, by comparing these correspondences to a DMPC/cholesterol phase diagram. Ternary systems of BSM/cholesterol/POPC were also studied, and it was found that the phase state of these systems is almost independent of temperature between 14-50^o C. Therefore, the ratio of liquid-ordered to liquid-disordered phases present in each sample was equal to the ratio of BSM/cholesterol to POPC.