Darabedian Narek, Pratt Matthew R
Department of Chemistry, University of Southern California, Los Angeles, CA, United States.
Department of Chemistry, University of Southern California, Los Angeles, CA, United States; Department of Biological Sciences, University of Southern California, Los Angeles, CA, United States.
Methods Enzymol. 2019;622:293-307. doi: 10.1016/bs.mie.2019.02.017. Epub 2019 Mar 6.
O-GlcNAcylation is a widespread posttranslational modification of intracellular proteins. Phenotypic and genetic experiments have established key roles for O-GlcNAc in development, mammalian cell survival, and several human diseases. However, the underlying mechanisms by which this modification alters biological pathways are still being discovered. An important part of this discovery process is the discovery of O-GlcNAcylated proteins, where chemical approaches have been particularly powerful. Here we describe how to combine one of these approaches, metabolic chemical reporters (MCRs), with bioorthogonal chemistry and Western blotting to identify potentially O-GlcNAcylated proteins.
O-连接的N-乙酰葡糖胺化（O-GlcNAcylation）是细胞内蛋白质广泛存在的一种翻译后修饰。表型和遗传学实验已证实O-GlcNAc在发育、哺乳动物细胞存活及多种人类疾病中发挥关键作用。然而，这种修饰改变生物途径的潜在机制仍在探索之中。这一发现过程的一个重要部分是对O-GlcNAc化蛋白质的发现，其中化学方法尤为有效。在此，我们描述如何将这些方法之一，即代谢化学报告分子（MCRs），与生物正交化学和蛋白质免疫印迹相结合，以鉴定潜在的O-GlcNAc化蛋白质。
