Evaluating Bilayer Mechanical Properties in Protein Reconstituted GUVs

Abstract

Biological membranes are complex systems where membrane proteins are surrounded by a bilayer composed of different types of lipids, with membranes from different organisms and organelles varying greatly in their composition and therefore physical properties. The mix of lipids in the membrane has additionally been shown to influence the activity, stability and conformation of many integral membrane proteins. In order to investigate the effect of incorporating protein on the mechanical properties of lipid bilayers we have reconstituted the Escherichia coli transporter lactose permease (LacY), a transmembrane protein representative of the major facilitator superfamily, into synthetic lipid vesicles. We observed directly the fluctuation of the lipid bilayer in these protein reconstituted GUVs, with sizes in the order of micrometers, by phase contrast microscopy. This allowed the determination of the bending rigidity, which characterizes the ability of membranes to bend under low stress, by fluctuation analysis. Changes in the bending rigidity parameter allowed us to get better insights into the effect of lipids and protein on the mechanical properties of GUVs in a quantitative fashion.