Ubiquitination is one of the most major post-translational modifications playing important role in regulation of intra-cellular proteins' stability, degradation, localization and biological activity. However, these proteins are difficult to be detected due to their low abundance, short half-life. In this study, ubiquitin-binding domains (UBDs) were constructed to purify the ubiquitinated proteins from Hep3B cells. Ubiquitinated proteins and sites were detected by LC-MS/MS. A total of 1 900 potential ubiquitinated proteins were identified. Among them, 158 ubiquitinated sites were identified, belonging to 102 proteins. Bioinformatics analysis revealed that the enriched pathways of ubiquitinated proteins were closely related to tumor occurrence and development. The dysfunction of ubiquitin-proteasome has a high correlation with cell signaling and extracellular matrix changing in tumor cells.
泛素化修饰是细胞内最重要的翻译后修饰形式之一，对细胞内蛋白质的稳定、降解、定位以及生物活性的调节起到重要作用。但因其在细胞内丰度低、降解周期短等特点而很难被检测。本研究中，制备的泛素结合结构域蛋白 (Ubiquitin-binding domains，UBDs) 用于富集肝癌细胞系Hep3B 中的泛素化蛋白，并通过液相色谱-串联质谱联用的方法对富集的泛素化蛋白进行鉴定。实验共鉴定到1 900 个潜在的泛素化蛋白和158 个泛素化位点，这些被鉴定到的泛素化位点分属于102 个蛋白。生物信息学分析发现泛素化蛋白显著富集的相关通路与肿瘤的发生发展密切相关，此结果暗示肿瘤细胞内泛素化-蛋白酶体的失调与肿瘤细胞的信号传导及细胞外基质的变化等具有较高的关联性。.
Keywords: Hep3B; ThUBDs; proteomics; ubiquitination.