Kakuda Shinako, Li Bingchen, London Erwin
Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, NY, United States.
Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, NY, United States.
Methods Enzymol. 2021;649:253-276. doi: 10.1016/bs.mie.2021.01.005. Epub 2021 Feb 18.
Studying the interaction of pore-forming toxins, including perfringolysin O (PFO), with lipid is crucial to understanding how they insert into membranes, assemble, and associate with membrane domains. In almost all past studies, symmetric lipid bilayers, i.e., bilayers having the same lipid composition in each monolayer (leaflet), have been used to study this process. However, practical methods to make asymmetric lipid vesicles have now been developed. These involve a cyclodextrin-catalyzed lipid exchange process in which the outer leaflet lipids are switched between two lipid vesicle populations with different lipid compositions. By use of alpha class cyclodextrins, it is practical to include a wide range of sterol concentrations in asymmetric vesicles. In this article, protocols for preparing asymmetric lipid vesicles are described, and to illustrate how they may be applied to studies of pore-forming toxin behavior, we summarize what has been learned about PFO conformation and its lipid interaction in symmetric and in asymmetric artificial lipid vesicles.
研究包括产气荚膜梭菌溶血素O（PFO）在内的成孔毒素与脂质的相互作用对于理解它们如何插入膜、组装以及与膜结构域结合至关重要。在几乎所有过去的研究中，对称脂质双层，即每个单层（小叶）具有相同脂质组成的双层，已被用于研究这一过程。然而，现在已经开发出制备不对称脂质囊泡的实用方法。这些方法涉及环糊精催化的脂质交换过程，其中外层小叶脂质在具有不同脂质组成的两个脂质囊泡群体之间切换。通过使用α类环糊精，可以在不对称囊泡中包含广泛的甾醇浓度。在本文中，描述了制备不对称脂质囊泡的方案，并且为了说明它们如何应用于成孔毒素行为的研究，我们总结了在对称和不对称人工脂质囊泡中关于PFO构象及其脂质相互作用所了解到的情况。
