Guest Paul C
Laboratory of Neuroproteomics, Department of Biochemistry and Tissue Biology, Institute of Biology, University of Campinas (UNICAMP), Cidade Universitária Zeferino Vaz, Rua Monteiro Lobato 255 F/01, 13083-862, Campinas, Brazil.
Adv Exp Med Biol. 2017;974:167-174. doi: 10.1007/978-3-319-52479-5_12.
Pulse radiolabelling of cells with radioactive amino acids such is a common method for investigating the biosynthetic rates of proteins. In this way, the abundance of newly synthesized proteins can be determined by several proteomic techniques including 2D gel electrophoresis (2DE). This chapter describes a protocol for labelling pancreatic islets with S-methionine in the presence of low and high concentrations of glucose, followed by subcellular fractionation enrichment of secretory granule proteins and analysis of the granule protein contents by 2DE. This demonstrated that the biosynthetic rates of most of the granule proteins are co-ordinately regulated in the presence of stimulatory glucose concentrations.
用放射性氨基酸对细胞进行脉冲放射性标记是研究蛋白质生物合成速率的常用方法。通过这种方式，可以用包括二维凝胶电泳（2DE）在内的几种蛋白质组学技术来测定新合成蛋白质的丰度。本章描述了一种在低浓度和高浓度葡萄糖存在下用³⁵S-蛋氨酸标记胰岛的方案，随后进行亚细胞分级分离以富集分泌颗粒蛋白，并通过二维凝胶电泳分析颗粒蛋白含量。这表明在刺激性葡萄糖浓度存在的情况下，大多数颗粒蛋白的生物合成速率受到协同调节。
