Lohse Martin J, Hoffmann Carsten, Nikolaev Viacheslav O, Vilardaga Jean-Pierre, Bünemann Moritz
Institute of Pharmacology and Toxicology, University of Würzburg, D-97078 Würzburg, Germany.
Adv Protein Chem. 2007;74:167-88. doi: 10.1016/S0065-3233(07)74005-6.
We describe and review methods for the kinetic analysis of G protein-coupled receptor (GPCR) activation and signaling that are based on optical methods. In particular, we describe the use of fluorescence resonance energy transfer (FRET) as a means of analyzing conformational changes within a single protein (for example a receptor) or between subunits of a protein complex (such as a G protein heterotrimer) and finally between distinct proteins (such as a receptor and a G protein). These methods allow the analysis of signaling kinetics in intact cells with proteins that retain their essential functional properties. They have produced a number of unexpected results: fast receptor activation kinetics in the millisecond range, similarly fast kinetics for receptor-G protein interactions, but much slower activation kinetics for G protein activation.
我们描述并综述了基于光学方法的G蛋白偶联受体（GPCR）激活和信号传导动力学分析方法。特别地，我们描述了利用荧光共振能量转移（FRET）来分析单个蛋白质（如受体）内或蛋白质复合物亚基间（如G蛋白异源三聚体）以及不同蛋白质间（如受体和G蛋白）的构象变化。这些方法能够在完整细胞中对保留其基本功能特性的蛋白质进行信号转导动力学分析。它们已经产生了许多意想不到的结果：毫秒级的快速受体激活动力学、受体 - G蛋白相互作用同样快速的动力学，但G蛋白激活的动力学要慢得多。
