Effect of physical constraints on the mechanisms of membrane fusion: bolaform lipid vesicles as model systems.

Bolaform lipid vesicles were used to study the effect of physical constraints on membrane fusion. In these vesicles the membrane is organized in a single monolayer, because of the presence of covalent bonds in its middle plane. Therefore, the formation of fusion intermediates is subject to higher energy barriers and greater geometrical constraints than is usual in bilayer membranes. Bolaform lipids were extracted from the thermophilic archaeon Sulfolobus solfataricus. These lipids can be divided into two classes, the monosubstituted molecules, in which one of the polar heads is glycerol, and the bisubstituted molecules, endowed with two complex polar heads. The fusion process in vesicles composed of different mixtures of monosubstituted/bisubstituted molecules was studied by means of fluorescence techniques. Ca2+ or poly(ethylene glycol) was employed as a fusogenic agent. We found that fusion of such constrained membranes is still possible, provided that molecules able to mediate a structural rearrangement of the membrane are present. This condition is fulfilled by monosubstituted molecules, which are able to partition the glycerol headgroup in the apolar moiety. In addition, the presence of traces (approximately 5%) of the monopolar compound diphytanylglycerol is an important factor for fusion to occur. On the contrary, vesicles formed by bisubstituted molecules are unable to fuse, irrespective of the fusogen employed.