Drunken Membranes: How does Ethanol Impact Fusion of Vesicles to Planar Lipid Bilayers?

Abstract

Previously we have shown that fusion of vesicles to a membrane is changed by lipid phase as altered by cholesterol and temperature (Lee et al. 2013, Chem Phys Lipids 166:45-54). Ethanol is also known to alter membrane phase behavior.To investigate the effect of ethanol on fusion rate, we used the Nystatin/Ergosterol (NYS/ERG) fusion assay. We measured fusions per minute using planar membranes containing PE/PC/CHL and vesicles containing PE/PC/PS/ERG. For our initial experiments we used high concentrations of ethanol added to either side of the bilayer. At 1% ethanol an increase was observed, and at 4% fusion rates increased 3 fold compared to control when added to the cis side of the bilayer (same side as vesicles). However, no significant increase was observed when added to the trans side. Similar results were observed with methanol. It also appears that the 2-3 fold increase in fusion rate occurs regardless of whether ethanol is added before or after vesicles.We hypothesize that this effect of ethanol is to alter lipid phase behavior of the vesicle membrane. To verify the effects of ethanol on membranes we utilized differential scanning calorimetry (nDSC). Solutions of KCl buffer and DPPC vesicles were analyzed with and without ethanol. The DPPC vesicles with ethanol were shown to have a transition state ∼0.5°C lower than without ethanol. This is consistent with our hypothesis that adding ethanol alters lipid fluidity and/or phase behavior.