CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai, China.
Methods Mol Biol. 2023;2564:299-315. doi: 10.1007/978-1-0716-2667-2_16.
Green fluorescent proteins (GFPs) have lightened up almost every aspect of biological research including protein sciences. In the field of membrane protein structural biology, GFPs have been used widely to monitor membrane protein localization, expression level, the purification process and yield, and the stability inside the cells and in the test tube. Of particular interest is the fluorescence-detector size-exclusion chromatography-based thermostability assay (FSEC-TS). By simple heating and FSEC, the generally applicable method allows rapid assessment of the thermostability of GFP-fused membrane proteins without purification. Here we describe the experimental details and some typical results for the FSEC-TS method.
绿色荧光蛋白（GFPs）已经点亮了几乎所有生物学研究领域，包括蛋白质科学。在膜蛋白结构生物学领域，GFPs 被广泛用于监测膜蛋白的定位、表达水平、纯化过程和产量，以及在细胞内和试管中的稳定性。特别有趣的是基于荧光探测大小排阻色谱的热稳定性测定法（FSEC-TS）。通过简单的加热和 FSEC，这种普遍适用的方法可以在不进行纯化的情况下快速评估 GFP 融合膜蛋白的热稳定性。本文描述了 FSEC-TS 方法的实验细节和一些典型结果。
