Department of Molecular Physiology and Biophysics, University of Fukui Faculty of Medical Sciences, 23-3 Matsuokashimoaizuki, Eiheiji-cho, Yoshida-gun, 910-1193, Fukui, Japan.
Sci Rep. 2017 Jul 31;7(1):6857. doi: 10.1038/s41598-017-07048-4.
In fluidic biomembranes, lipids and membrane proteins diffuse restlessly, and lipid compositions change steadily. To mimic dynamic behavior of the biomembranes, a method for introducing rapid changes in the constituents in the lipid bilayer was developed. In contact bubble bilayers (CBB), as a water-in-oil droplet bilayer system, the bilayer hydrophobic interior is contiguous with the bulk oil phase. Making use of this geometrical feature as an access route, hydrophobic substances were administered into the bilayer. Polytheonamide B, a cytotoxic hydrophobic peptide, was applied, and oriented incorporation and relevant single-channel current recordings were enabled. Nystatin was pre-loaded in the CBB, and sterol perfusion exhibited slow development of the macroscopic current. On the contrary, the reconstituted KcsA potassium channels immediately attenuate the channel activity when cholesterol was applied. This oil-phase route in the CBB allows rapid perfusion of hydrophobic substances around the bilayer-embedded channels during continuous recordings of channel currents.
在流体生物膜中，脂质和膜蛋白会不安分地扩散，脂质组成也会稳定地变化。为了模拟生物膜的动态行为，开发了一种在脂质双层中快速改变成分的方法。在接触气泡双层（CBB）中，作为水包油滴双层系统，双层疏水性内部与大块油相连续。利用这个几何特征作为进入途径，将疏水性物质施用到双层中。应用了多粘菌素 B，一种细胞毒性疏水性肽，并实现了定向掺入和相关的单通道电流记录。制霉菌素预先加载在 CBB 中，甾醇灌注显示出宏观电流的缓慢发展。相反，当胆固醇施加时，再构成的 KcsA 钾通道立即减弱通道活性。CBB 中的油相途径允许在连续记录通道电流时，在嵌入双层的通道周围快速灌注疏水性物质。
