Transbilayer distribution of phosphatidylethanolamine in large and small unilamellar vesicles.

Abstract

There is much evidence which strongly suggests that most constituents of biological membranes display a transbilayer compositional asymmetry. The tendency of binary mixtures of phospholipids to form compositionally asymmetric bilayers spontaneously has been studied extensively. In small unilamellar vesicles, most mixtures of phospholipids with different head groups have been reported to be nonrandomly arranged across the bilayer. In this study, the influence of the radius of curvature on the transbilayer phospholipid distribution has been investigated. The distribution of egg phosphatidylethanolamine in large unilamellar vesicles comprised of egg phosphatidylethanolamine and egg was determined by 2,4,6-trinitrobenzenesulfonic acid labeling. These large vesicles were obtained by modifying the ethanol injection procedure originally described by Batzri & Korn (1973) [Batzri, S., & Korn, E. D. (1973) Biochim. Biophys. Acta 298, 1015] by using a slow injection rate. After injection, the ethanol was removed by molecular sieve chromatography and the vesicle dispersion centrifuged. This results in a population of large, homogeneous, and unilamellar vesicles as determined by molecular sieve chromatography, 32P NMR, and electron microscopy. The phosphatidylethanolamine component in unilamellar vesicles of this type is equally distributed between the two monolayers. In contrast, phosphatidylethanolamine in small unilamellar vesicles is known to be preferentially localized in the outer monolayer at low phosphatidylethanolamine concentrations and in the inner monolayer at high phosphatidylethanolamine concentrations. These results suggest that while phospholipids may form asymmetric bilayers spontaneously in highly curved regions of biological membranes, other factors must be responsible for the generalized phospholipid asymmetry seen in these systems.