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Fusion of lipid bilayers induced by synthetic lipids and peptides

University of British Columbia

The fusion-inducing properties of three different synthetic lipid derivatives were investigated in model membranes. The results are discussed in terms of membrane destabilization caused by the formation of non-bilayer lipid structures following charge neutralization of the fusogenic components. First, studies on the fusion of large unilamellar vesicles (LUVs) composed of equimolar 1,2-dioleoyl-OT-phosphatidylethanolamine (DOPE) and the synthetic cationic lipid N,Ndimethyl- A/,Af-di-9-octadecenylammonium chloride (DODAC) with target membranes of varying lipid composition are presented. Membrane fusion was promoted by increased negative surface charge and, for liquid-crystalline lipids, by increased acyl chain unsaturation in target liposomes. However, the presence of disaturated phospholipids promoted fusion below the gel-to-liquid-crystalline transition temperature, an effect which was eliminated by the addition of cholesterol. It was also shown that DOPE/DODAC (1:1) LUVs fused with erythrocyte membranes and that this fusion was blocked by the presence of serum. Next, fusion of model lipid bilayers containing synthetic aminolipids and regulation of this fusion by inducing transbilayer asymmetry of these aminolipids via imposed pH gradients is discussed. Liposomes consisting of 5 mol% ±-l,2-dioleoyl-3-N,N-dimethylaminopropane (AL1) in egg phosphatidylcholine/DOPE/cholesterol (EPC/DOPE/Chol, 35:20:45) did not fuse at pH 4.0 or upon increasing the external pH of these vesicles to 7.5, which resulted in rapid transport of AL1 to the inner monolayer. However, dissipation of the imposed pH gradient led to redistribution of AL1 to the outer monolayer at pH 7.5 and caused liposomal fusion. The effect of varying the hydrocarbon structure of AL1 on the rate of fusion was demonstrated with five synthetic analogues. Finally, fusion of lipid bilayers induced at low pH by a synthetic lipopeptide (Lipo- AcE4K) derived from the fusion peptide of influenza hemagglutinin is discussed. The secondary structure of Lipo-AcE4K was not affected by pH, but increased membrane penetration was indicated by tryptophan fluorescence. Lipo-AcE4K formed stable liposomes with l-palmitoyl-2-oleoyl-jn-phosphatidylcholine (POPC) and EPC/Chol (55:45) at pH 7.5 but induced fusion of these vesicles at mildly acidic pH. Membrane destabilization increased with increasing lipopeptide concentrations, decreasing pH, inclusion of cholesterol, and incorporation of lipopeptide into the liposomal inner monolayer. Fusion of liposomes bearing Lipo-AcE4K with erythrocyte membranes was also demonstrated.

Thesis/Dissertation

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2009-03-19

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http://hdl.handle.net/2429/6258

Biochemistry and Molecular Biology

University of British Columbia

UBCV

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