Fluidity Parameters of Lipid Regions Determined by Fluorescence Polarization

Lipid structural order parameters (reciprocal of fluidity) in biomembranes derived from steady-state fluorescence polarization measurements.

Penetration and enzymatic barriers to peptide and protein absorption

C) Means to enhance penetration

Publisher Summary This chapter discusses the dynamics and organization of lipid assemblies derives from studies of model lipid membranes of well-defined chemical composition. However, because of the heterogeneity in both the lipid composition and in the acyl chains of single phospholipids, which is inherent in biological membranes, extrapolations from the model system can only be qualitative in nature. The current notion of lipid organization in biological membranes is of a complex and heterogeneous structure. In areas where the protein to lipid ratio is small, the membrane presumably behaves in accord with the fluid mosaic model, where the lipids form a fluid pool in which proteins are embedded to different extents. The modes of diffusion of the proteins as asserted by this model are mostly passive and are, therefore, determined to a large extent by the fluidity of the lipid matrix. In other membrane areas, the protein to lipid ratio may be greater and the structure may approach a loosely bound network of proteins among which lakes of lipids are spread.

Fluidity of cell membranes--current concepts and trends.

Rat intestinal microvillus membranes and lipid extracts prepared from them have been studied by fluorescence polarization with three lipid-soluble fluorophores: diphenylhexatriene, retinol, and anthroyl-stearate. The degree of fluorescence polarization of diphenylhexatriene, which provides an index of the "microviscosity" of the lipid regions of the membrane, is exceptionally high in microvillus membranes, the highest yet reported in normal biological membranes. Both the membrane proteins and lipids were found to contribute to the high values. With each of the three probes the polarization values are higher in ileal microvillus membranes as compared to membranes from proximal intestinal segments. Temperature-dependence studies of the fluorescence polarization of diphenylhexatriene and anthroylstearate demonstrate a phase transition in microvillus membranes and in liposomes prepared from their lipid extracts at approximately 26+/-2 degrees C. Ambient pH influences markedly the diphenylhexatriene fluorescence polarization in microvillus membranes but has little effect on that of human erythrocyte ghost membranes. The "microviscosity" of jejunal microvillus membranes is maximal at pH 6.5-7.0 and decreases as much as 50% at pH 3.0, an effect which depends largely upon the membrane proteins. Addition of calcium ions to suspensions of microvillus membranes increases the fluorescence polarization of retinol and anthroyl-stearate, but not that of diphenyl-hexatriene. This confirms the localization of the last compound to the hydrophobic interior of the membrane, relatively distant from the hydrophilic head groups of the polar lipids. Microvillus membrane proteins solubilized with Triton X-100 give relatively high fluorescence polarization and intensity values with retinol, suggesting the presence of binding proteins which could play a role in the normal absorptive mechanism for the vitamin.

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M Shinitzky

Papers

Fluorescence polarization studies of rat intestinal microvillus membranes.