Interaction of siglec protein with glycolipids in a lipid bilayer deposited on a gold electrode surface

Unsymmetric three component lipid bilayers containing 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC), cholesterol and a ganglioside were prepared using the Langmuir–Blodgett–Langmuir–Schaefer (LB–LS) technique on the surface of a polycrystalline gold electrode. The lipid layer contains the ganglioside only in the outer layer, mimicking a natural cell membrane. Gangliosides are glycolipids containing at least one sialic acid residue and are proposed to be a binding partner for siglec proteins. Interaction between Siglec-4, the myelin-associated glycoprotein (MAG) and two gangliosides in lipid bilayers was investigated. MAG protein binds only to the sialic acid residue located at the terminal position in the oligosaccharide head group of a lipid molecule. In this study two gangliosides, the GD1a having one terminal sialic acid residue and the GM1 not possessing a terminal sialic acid residue in the polar head group were used. Under physiological conditions a lipid membrane is constantly exposed to high electric fields. Three component lipid bilayers transferred by the LB–LS method are characterized by capacitance minimum equal to 2.3 and 5.0 μF cm−2 in the film containing the GM1 and GD1a, respectively. The lipid bilayer desorbs form the electrode surface at E < −0.8 V vs. Ag|AgCl. The potential driven adsorption–desorption process is irreversible and leads to an increase in the capacitance of the adsorbed lipid films. The lipid films undergo some irreversible changes which may lead to a flip–flop of bilayer fragments, formation of micelles on the electrode surface or decrease in surface coverage. Exposure of the bilayer to the protein solution stabilizes lipid films adsorbed directly on the electrode surface. The potential induced adsorption of the bilayer containing the GD1a in the presence of the protein hinders structural changes taking place in the lipid film exposed to changing electric fields. At the molecular level the binding of the protein to the GD1a introduces structural changes not only in the glycolipid molecule, but in the whole membrane. In the polar head group of the GD1a amide groups in sialic acid residues lose some water of hydration upon interaction with the protein. The hydrocarbon chains of the GD1a exhibit ordering with respect to the surface normal. Reorientations in the ganglioside molecule may lead to changes in the phosphatidylcholine, main lipid component of the bilayer. In the DMPC molecule the dehydration of the ester group with simultaneous change in tilt of the acyl chains were observed.