Supported Lipid Bilayers on Biocompatible Polysaccharide Multilayers

Abstract

Solid-supported phospholipid bilayers are widely used as models to probe the function of cellular membranes and as platforms for membrane protein biosensing applications. Although supported lipid bilayers have a number of advantages in terms of ease of fabrication and characterization, the proximity of the membrane to the solid support is a limitation for applications that involve incorporation of integral membrane proteins. This limitation can be overcome by formation of supported lipid bilayers on soft polymer cushions. We have prepared biocompatible polymer cushions by the layer-by-layer assembly of two polysaccharide polyelectrolytes, chitosan (CHI) and hyaluronic acid (HA), on glass substrates. (CHI/HA)5 films were characterized by atomic force microscopy, giving an average thickness of 57 and roughness of 25 nm in aqueous solution at pH 6.5. Formation of zwitterionic lipid bilayers by the vesicle fusion method was attempted using DOPC vesicles at pH 4 and pH 6.5 on (CHI/HA)5 films. At higher pH adsorbed lipids had low mobility and large immobile lipid fractions; a combination of fluorescence and AFM indicated that this was due to the formation of heterogeneous membranes with defects and pinned lipids, rather than to a layer of surface adsorbed vesicles. By contrast, more uniform bilayers with mobile lipids were produced at pH 4. Fluorescence recovery after photobleaching gave diffusion coefficients that were similar to those for bilayers on PEG cushions and considerably higher than those measured on other polyelectrolyte films. The formation of bilayers containing reconstituted integral membrane protein was compared on PEM and PEG films. These results demonstrate that polysaccharides provide a useful alternative to other polymer cushions, particularly for applications where biocompatibility is important.