Protein-Lipid Interaction and Domain Formation in Asymmetric Membranes

Abstract

The lateral organization of lipids and receptors in cellular membranes is connected to a multitude of biological and pathological processes. The role of sphingolipid- and cholesterol-rich lipid “rafts” as membrane domains that control protein-protein interaction has recently attracted much attention. While the model membrane approach has yielded fundamental insights into cell membrane structure and function, the majority of these studies did not take into account the very important asymmetry between the composition of the inner and the outer leaflets of cell membranes. Therefore, certain key questions remain still unanswered: for example, how is the partition of membrane components in raft domains affected by the presence of non-raft lipids in the inner leaflet? To answer questions of this type we have developed a new method based upon cyclodextrin-induced lipid exchange to create giant unilamellar vesicles (GUVs) featuring the same type of asymmetry encountered in biological membranes. These vesicles can be produced with a very large yield, while avoiding use of organic solvents. The asymmetry of the bilayer can be confirmed by leaflet-targeted Fluorescence Correlation Spectroscopy (FCS). The use of asymmetric GUVs as novel model bilayers should allow the investigation of the principles underlying communication between raft-like domains in opposite leaflets of the bilayer, as well as investigation of the origin of lipid and membrane protein affinity for rafts.