UNASSIGNED: Droplet interface bilayers (DIBs) serve as a convenient platform to study interactions between synthetic lipid membranes and proteins. However, a majority of DIBs have been assembled using a single lipid type, @entity68722 ( @entity68723 ). The work described herein establishes a new method to assemble DIBs using total lipid extract from @entity526 (eTLE); it is found that incubating oil-submerged aqueous droplets containing eTLE liposomes at a temperature above the gel-fluid phase transition temperature (Tg) promotes monolayer self-assembly that does not occur below Tg. Once monolayers are properly assembled via heating, droplets can be directly connected or cooled below Tg and then connected to initiate bilayer formation. This outcome contrasts immediate droplet coalescence observed upon contact between nonheated eTLE-infused droplets. Specific capacitance measurements confirm that the interface between droplets containing eTLE lipids is a lipid bilayer with thickness of 29.6    at 25   C in @entity2291 . We observe that bilayers formed from eTLE or @entity68723 survive cooling and heating between 25 and 50   C and demonstrate gigaohm (G  ) membrane resistances at all temperatures tested. Additionally, we study the insertion of @entity37114 peptides into both eTLE and @entity68723 membranes to understand how lipid composition, temperature, and membrane phase influence ion channel formation. Like in @entity68723 bilayers, @entity37114 peptides in eTLE exhibit discrete, voltage-dependent gating characterized by multiple open channel conductance levels, though at significantly lower applied voltages. Cyclic voltammetry measurements of macroscopic channel currents confirm that the voltage-dependent conductance of @entity37114 channels in eTLE bilayers occurs at lower voltages than in @entity68723 bilayers at equivalent peptide concentrations. This result suggests that eTLE membranes, via composition, fluidity, or the presence of subdomains, offer an environment that enhances @entity37114 insertion. For both membrane compositions, increasing temperature reduces the lifetimes of single channel gating events and increases the voltage required to cause an exponential increase in channel current. However, the fact that @entity37114 insertion in eTLE exhibits significantly greater sensitivity to temperature changes through its Tg suggests that membrane phase plays an important role in channel formation. These effects are much less severe in @entity68723 , where heating from 25 to 50   C does not induce a phase change. The described technique for heating-assisted monolayer formation permits the use of other high transition temperature lipids in aqueous droplets for DIB formation, thereby increasing the types of lipids that can be considered for assembling model membranes.

Heating-Enabled Formation of Droplet Interface Bilayers Using XXXX Total Lipid Extract.

@entity68723 :: ('MESH:C060859', 'Chemical') :: ['DPhPC'] @entity37114 :: ('MESH:D000408', 'Chemical') :: ['alamethicin'] @entity2291 :: ('MESH:C007932', 'Chemical') :: ['hexadecane'] @entity68722 :: ('MESH:C021016', 'Chemical') :: ['diphytanoylphosphatidylcholine'] @entity526 :: ('562', 'Species') :: ['Escherichia coli']
@entity526 :: (562,Species) :: ['Escherichia coli']
