Nandi Animesh, Sprung Robert, Barma Deb K, Zhao Yingxin, Kim Sung Chan, Falck John R, Zhao Yingming
Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9038, USA.
Anal Chem. 2006 Jan 15;78(2):452-8. doi: 10.1021/ac051207j.
The O-linked N-acetylglucosamine (O-GlcNAc) modification of serine/threonine residues is an abundant posttranslational modification present in cytosolic and nuclear proteins. The functions and subproteome of O-GlcNAc modification remain largely undefined. Here we report the application of the tagging-via-substrate (TAS) approach for global identification of O-GlcNAc-modified proteins. The TAS method utilizes an O-GlcNAc azide analogue for metabolic labeling of O-GlcNAc-modified proteins, which can be chemoselectively conjugated for detection and enrichment of the proteins for proteomics studies. Our study led to the identification of 199 putative O-GlcNAc-modified proteins from HeLa cells, among which 23 were confirmed using reciprocal immunoprecipitation. Functional classification shows that proteins with diverse functions are modified by O-GlcNAc, implying that O-GlcNAc might be involved in the regulation of multiple cellular pathways.
丝氨酸/苏氨酸残基的O-连接的N-乙酰葡糖胺（O-GlcNAc）修饰是一种存在于胞质和核蛋白中的丰富的翻译后修饰。O-GlcNAc修饰的功能和亚蛋白质组在很大程度上仍不明确。在此，我们报告了通过底物标记（TAS）方法用于全面鉴定O-GlcNAc修饰蛋白。TAS方法利用一种O-GlcNAc叠氮类似物对O-GlcNAc修饰蛋白进行代谢标记，其可进行化学选择性共轭以检测和富集用于蛋白质组学研究的蛋白。我们的研究从HeLa细胞中鉴定出199种推定的O-GlcNAc修饰蛋白，其中23种通过相互免疫沉淀得到证实。功能分类表明具有多种功能的蛋白被O-GlcNAc修饰，这意味着O-GlcNAc可能参与多种细胞途径的调控。
