Reconstitution and Electrophysiological Characterization of Ion Channels in Lipid Bilayers.
Reconstitution and Electrophysiological Characterization of Ion Channels in Lipid Bilayers.
复制标题
DOI：
10.1002/cpph.37
复制
发表时间：
2018-06-01
影响因子：
--
通讯作者：
Calloe, Kirstine
中科院分区：
文献类型：
--
作者：
Klaerke, Dan A;Tejada, Maria de Los Angeles;Calloe, Kirstine
洗涤剂增溶的纯化离子通道可以重组为脂质双层，用于电生理分析。传统上，离子通道被插入囊泡中，然后与脂类溶解在疏水溶剂(如正庚烷)中形成的平面“黑色脂膜”融合在一起。本文提供了一种将纯化的离子通道蛋白重组为巨大的单层囊泡(GUV)的逐步指南。这一过程导致了可用于平面双层形成和单通道电流的电生理表征的蛋白质脂质体的形成。通过使用预制的GUV，有可能省略膜溶剂。与传统制剂相比，由GUV形成的脂双层提供了更接近天然细胞膜的环境。还描述了一种替代方案，该方案需要从将洗涤剂中的蛋白质添加到其上的GUV产生平面脂双层。2018年John Wiley&Sons，Inc.
Detergent-solubilized purified ion channels can be reconstituted into lipid bilayers for electrophysiological analysis. Traditionally, ion channels were inserted into vesicles and subsequently fused with planar "black lipid membranes" formed from lipids dissolved in a hydrophobic solvent such as decane. Provided in this article is a step-by-step guide to reconstitute purified ion channel proteins into giant unilamellar vesicles (GUVs). This procedure results in the formation of proteoliposomes that can be used for planar bilayer formation and electrophysiological characterization of single-channel currents. By using preformed GUVs it is possible to omit the membrane solvent. Compared to traditional preparations, the lipid bilayers formed from GUVs provide an environment that more closely resembles the native cell membrane. Also described is an alternate protocol that entails the production of planar lipid bilayers from GUVs onto which proteins in detergent are added. © 2018 by John Wiley & Sons, Inc.