O-linked β-N-acetylglucosamine (O-GlcNAc) is an essential regulatory post-translational modification of thousands of nuclear, cytoplasmic and mitochondrial proteins. O-GlcNAc is dynamically added and removed from proteins by the O-GlcNAc transferase and the O-GlcNAcase (OGA), respectively. Dysregulation of O-GlcNAc-cycling is implicated in the etiology of numerous diseases including tumorigenesis, metabolic dysfunction and neurodegeneration. To facilitate studies focused on the role of O-GlcNAc and OGA in disease, we sought to identify commercially available antibodies that enable the enrichment of full-length OGA (fOGA) from lysates of mouse and human origin. Here, we report that antibodies from Abcam and Bethyl Laboratories can be used to immunoprecipitate OGA to near-saturation from human and mouse cell lysates. However, western blotting analysis indicates that both antibodies, as well as three noncommercially available antibodies (345, 346, 352), detect fOGA and numerous cross-reacting proteins. These nonspecific signals migrate similarly to fOGA and are detected robustly, suggesting that the use of appropriate controls is essential to avoid the misidentification of OGA.

中文翻译：

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用于检测和丰富人和小鼠O- GlcNAcase的工具的表征

O-连接的β- N-乙酰氨基葡萄糖（O- GlcNAc）是成千上万的核，细胞质和线粒体蛋白质的重要​​调控翻译后修饰。O- GlcNAc分别通过O- GlcNAc转移酶和O- GlcNAcase（OGA）从蛋白质中动态添加和去除。O - GlcNAc-循环的失调与多种疾病的病因有关，包括肿瘤发生，代谢功能障碍和神经变性。为了促进针对O-的作用的研究GlcNAc和OGA在疾病中，我们试图鉴定可从小鼠和人类来源的裂解物中富集全长OGA（fOGA）的市售抗体。在这里，我们报道来自Abcam和Bethyl Laboratories的抗体可用于从人和小鼠细胞裂解物中免疫沉淀OGA至近乎饱和。然而，蛋白质印迹分析表明，两种抗体以及三种非商业可用的抗体（345、346、352）都可以检测到fOGA和许多交叉反应蛋白。这些非特异性信号以类似于fOGA的方式迁移，并被可靠地检测到，这表明使用适当的控件对于避免OGA的错误识别至关重要。