Institute of Biochemistry, Vilnius University, Vilnius, Lithuania.
Langmuir. 2013 Apr 2;29(13):4320-7. doi: 10.1021/la304613a. Epub 2013 Mar 18.
Phosphatidylcholine and cholesterol exchange between vesicles and planar tethered bilayer lipid membranes (tBLMs) was demonstrated from electrochemical impedance spectroscopy (EIS), fluorescence microscopy (FM), and neutron reflectometry (NR) data. Cholesterol is incorporated into the tBLMs, as determined by the functional reconstitution of the pore forming toxin α-hemolysin (EIS data), attaining cholesterol concentrations nearly equal to that in the donor vesicles. Using fluorescently labeled lipids and cholesterol, FM indicates that the vesicle-tBLM exchange is homogeneous for the lipids but not for cholesterol. NR data with perdeuterated lipids indicates lipid exchange asymmetry with two lipids exchanged in the outer leaflet for every lipid in the inner leaflet. NR and EIS data further show different exchange rates for cholesterol (t1/2 < 60 min) and phosphatidylcholine (t1/2 > 4 h). This work lays the foundation for the preparation of robust, lower defect, more biologically relevant tBLMs by essentially combining the two methods of tBLM formation-rapid solvent exchange and vesicle fusion.
通过电化学阻抗谱（EIS）、荧光显微镜（FM）和中子反射测量（NR）数据，证明了磷脂酰胆碱和胆固醇在囊泡和平面连接双层脂质膜（tBLM）之间的交换。通过孔形成毒素α-溶血素的功能重建（EIS 数据）确定了胆固醇掺入 tBLM，达到了与供体囊泡中胆固醇浓度几乎相等的水平。使用荧光标记的脂质和胆固醇，FM 表明脂质在囊泡-tBLM 交换中是均匀的，但胆固醇不是。具有氘化脂质的 NR 数据表明脂质交换具有不对称性，在外层中交换两个脂质，内层中交换一个脂质。NR 和 EIS 数据还表明胆固醇（t1/2 < 60 min）和磷脂酰胆碱（t1/2 > 4 h）的交换速率不同。这项工作为制备更稳健、缺陷更少、更具生物学相关性的 tBLM 奠定了基础，本质上是将两种 tBLM 形成方法——快速溶剂交换和囊泡融合结合起来。
