Prasannan Praseetha, Saravanan Rathi, Verma Navin Kumar
Lymphocyte Signalling Research Laboratory, Lee Kong Chian School of Medicine, Nanyang Technological University Singapore, Singapore, Singapore.
Lee Kong Chian School of Medicine, Nanyang Technological University Singapore, Singapore, Singapore.
Methods Mol Biol. 2019;1930:83-90. doi: 10.1007/978-1-4939-9036-8_11.
Protein phosphorylation plays a key role in intracellular signal transduction and regulates diverse cellular functions. This posttranslational modification of proteins occurs dynamically and reversibly and only a small fraction of the total proteins is phosphorylated at any given time depending on the cell types and their functioning. Thus, a relatively low abundance of phosphorylated proteins is present in specific cells under certain conditions and hence it becomes problematic to detect these proteins and their analysis. In particular, phosphoproteomic analysis of rapidly migrating T-lymphocytes is always challenging. In order to analyze phosphoproteins in motile T-cells using techniques such as polyacrylamide gel electrophoresis and mass spectrometry, it is often important to enrich the phosphorylated forms in the cellular lysates. In this chapter, we describe a simple method to enrich phosphoproteins that can be used for protein analysis in motile T-cells.
蛋白质磷酸化在细胞内信号转导中起关键作用，并调节多种细胞功能。这种蛋白质的翻译后修饰动态且可逆地发生，在任何给定时间，根据细胞类型及其功能，只有一小部分总蛋白质被磷酸化。因此，在特定条件下，特定细胞中磷酸化蛋白质的丰度相对较低，因此检测这些蛋白质及其分析变得很困难。特别是，对快速迁移的T淋巴细胞进行磷酸化蛋白质组分析一直具有挑战性。为了使用聚丙烯酰胺凝胶电泳和质谱等技术分析运动性T细胞中的磷酸化蛋白质，通常重要的是在细胞裂解物中富集磷酸化形式。在本章中，我们描述了一种富集磷酸化蛋白质的简单方法，该方法可用于运动性T细胞中的蛋白质分析。
