Dual inhibitory effect of gangliosides on phospholipase C-promoted fusion of lipidic vesicles

The effect of a variety of gangliosides has been tested on the phospholipase C-induced fusion of large unilamellar vesicles. Bilayer composition was phosphatidylcholine:phosphatidylethanolamine:cholesterol (2:1:1 mole ratio) plus the appropriate amounts of glycosphingolipids. Enzyme phosphohydrolase activity, vesicle aggregation, mixing of bilayer lipids and mixing of liposomal aqueous contents were separately assayed. Small amounts (<1 mol %) of gangliosides in the lipid bilayer produce a significant inhibition of the above processes. The inhibitory effect of gangliosides increases with the size of the oligosaccharide chain in the polar head group. Inhibition depends in a nonlinear manner on the ganglioside proportion, and is complete at similar to 5 mol %. Inhibition is not due to ganglioside-dependent changes in vesicle curvature or size. Ganglioside inhibition of vesicle fusion is due to two different effects: inhibition of phospholipase C activity and stabilization of the lipid lamellar phase. Enzyme inhibition leads to a parallel decrease of vesicle aggregation and lipid mixing rates. Mixing of aqueous contents, though, is depressed beyond the enzyme inhibition levels. This is explained in terms of the fusion pore requiring a local destabilization of the lipid bilayer, the lamellar structure being stabilized by gangliosides. P-31-NMR and DSC experiments confirm the inhibitory effect of gangliosides in various lamellar-to-nonlamellar transitions.