Department of Chemistry and Molecular Biology, Gothenburg University, Box 462, 405 30, Göteborg, Sweden.
Sci Rep. 2021 Sep 28;11(1):19232. doi: 10.1038/s41598-021-98810-2.
Interactions between membrane proteins within a cellular environment are crucial for all living cells. Robust methods to screen and analyse membrane protein complexes are essential to shed light on the molecular mechanism of membrane protein interactions. Most methods for detecting protein:protein interactions (PPIs) have been developed to target the interactions of soluble proteins. Bimolecular fluorescence complementation (BiFC) assays allow the formation of complexes involving PPI partners to be visualized in vivo, irrespective of whether or not these interactions are between soluble or membrane proteins. In this study, we report the development of a screening approach which utilizes BiFC and applies flow cytometry to characterize membrane protein interaction partners in the host Saccharomyces cerevisiae. These data allow constructive complexes to be discriminated with statistical confidence from random interactions and potentially allows an efficient screen for PPIs in vivo within a high-throughput setup.
在细胞环境中，膜蛋白之间的相互作用对于所有活细胞都是至关重要的。开发强大的方法来筛选和分析膜蛋白复合物对于揭示膜蛋白相互作用的分子机制至关重要。大多数用于检测蛋白质-蛋白质相互作用（PPIs）的方法都是针对可溶性蛋白相互作用开发的。双分子荧光互补（BiFC）测定法允许在体内可视化涉及 PPI 伙伴的复合物的形成，而不管这些相互作用是在可溶性蛋白还是膜蛋白之间。在这项研究中，我们报告了一种筛选方法的开发，该方法利用 BiFC 并应用流式细胞术来表征宿主酿酒酵母中的膜蛋白相互作用伙伴。这些数据允许以统计置信度区分有建设性的复合物与随机相互作用，并可能允许在高通量设置中有效地筛选体内的 PPIs。
