Unitat de Farmacologia, Departament de Patologia i Terapèutica Experimental, Facultat de Medicina, IDIBELL-Universitat de Barcelona, 08907 Barcelona, Spain.
Trends Biotechnol. 2010 Aug;28(8):407-15. doi: 10.1016/j.tibtech.2010.05.002. Epub 2010 Jun 9.
Spatiotemporal characterization of protein-protein interactions (PPIs) is essential in determining the molecular mechanisms of intracellular signaling processes. In this review, we discuss how new methodological strategies derived from non-invasive fluorescence- and luminescence-based approaches (FRET, BRET, BiFC and BiLC), when applied to the study of G protein-coupled receptor (GPCR) oligomerization, can be used to detect specific PPIs in live cells. These technologies alone or in concert with complementary methods (SRET, BRET or BiFC, and SNAP-tag or TR-FRET) can be extremely powerful approaches for PPI visualization, even between more than two proteins. Here we provide a comprehensive update on all the biotechnological aspects, including the strengths and weaknesses, of new fluorescence- and luminescence-based methodologies, with a specific focus on their application for studying PPIs.
蛋白质-蛋白质相互作用(PPIs)的时空特征对于确定细胞内信号转导过程的分子机制至关重要。在这篇综述中,我们讨论了如何将源自非侵入性荧光和发光方法(FRET、BRET、BiFC 和 BiLC)的新方法策略应用于 G 蛋白偶联受体(GPCR)寡聚化的研究,以在活细胞中检测特定的 PPIs。这些技术单独或与互补方法(SRET、BRET 或 BiFC 和 SNAP-tag 或 TR-FRET)结合使用,可以成为 PPI 可视化的极其强大的方法,即使是在两个以上的蛋白质之间。在这里,我们全面更新了所有基于荧光和发光的新方法学的生物技术方面,包括它们的优缺点,并特别关注它们在研究 PPIs 中的应用。