Kobayashi Fuminori, Nishiuchi Takumi, Takaki Kento, Konno Hiroki
Graduate School of Natural Science & Technology, Kanazawa University, Kanazawa 920-1192, Japan.
Institute for Gene Research Center, Bio-AFM Frontier Research Center, Kanazawa University, Kanazawa 920-1192, Japan.
Biochem Biophys Res Commun. 2018 Feb 5;496(2):686-692. doi: 10.1016/j.bbrc.2017.12.076. Epub 2017 Dec 27.
Ubiquitination of target proteins is accomplished by isopeptide bond formation between the carboxy group of the C-terminal glycine (Gly) residue of ubiquitin (Ub) and the ɛ-amino group of lysine (Lys) on the target proteins. The formation of an isopeptide bond between Ubs that gives rise to a poly-Ub chain on the target proteins and the types of poly-Ub chains formed depend on which of the seven Lys residues or N-terminal methionine (Met) residue on Ub is used for chain elongation. To understand the linkage specificity mechanism of Ub chains on E3, the previous study established an assay to monitor the formation of a free diubiquitin chain (Ub chain synthesis assay) by HECT type E3 ligase. In this study, we investigated Ub chain specificity using E6AP HECT domain. We here demonstrate the importance of the N-terminal domain of full length E6AP for Ub chain specificity.
靶蛋白的泛素化是通过泛素（Ub）C末端甘氨酸（Gly）残基的羧基与靶蛋白上赖氨酸（Lys）的ε-氨基之间形成异肽键来实现的。在靶蛋白上产生多聚泛素链的泛素之间异肽键的形成以及所形成的多聚泛素链的类型取决于泛素上七个赖氨酸残基或N末端甲硫氨酸（Met）残基中的哪一个用于链延伸。为了了解E3上泛素链的连接特异性机制，之前的研究建立了一种检测方法，以监测HECT型E3连接酶形成游离双泛素链（泛素链合成检测）。在本研究中，我们使用E6AP HECT结构域研究了泛素链特异性。我们在此证明全长E6AP的N末端结构域对泛素链特异性的重要性。
