Vesicle Budding Induced by the Asymmetric Membrane Insertion of a Surfactant is Limited by an Osmotic Barrier

Abstract

Due to medical, technical and economical importance, substantial efforts have been made to further understand interactions of surfactants and lipid-bilayers. While approaches such as the well known three-stage-model - describing solubilization of lipid bilayers by surfactants - serve their purpose, complex systems escape its applicability. One such case is where equilibration of surfactant across the bilayer does not occur rapidly after addition from the outside, i.e. surfactant flip does not occur on the observed timescale. In this scenario, stress is induced by non-matching surface area requirements of the inner and outer leaflet. Several mechanisms for the release of this stress have been proposed. 12:0 Lyso-PC and POPC liposomes represent such a system. Here, asymmetry stress stops further lysolipid uptake beyond a certain content and gives rise to originally largely pure lysolipid micelles that equilibrate with the liposomes over days only (“staying out” scenario). However, part of the stress is also relaxed by a limited number of very small exovesicle budding off from the mother vesicle. Here, we present a first quantification of the extent of budding that accompanies “staying out”. This is achieved by separating the budded exovesicles from the mother vesicles using Asymmetric Flow-Field-Flow Fractionation (AF4). We studied the correlation of the extent of budding with osmotic pressure gradients, which affect the sphericity of the liposomes before addition of lysolipid. It suggests that fast budding from liposomes in buffer proceeds under constant entrapped volume conditions and, hence, stops as the mother vesicle becomes spherical. Further budding is slow, probably limited by ion leakage across the membrane. The detailed understanding of the budding processes is valuable both for technical applications and for deeper insight into natural budding-related phenomena such as exo- and endocytosis.