A ubiquitin ligase transfers preformed polyubiquitin chains from a conjugating enzyme to a substrate
A ubiquitin ligase transfers preformed polyubiquitin chains from a conjugating enzyme to a substrate
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DOI：
10.1038/nature05542
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发表时间：
2007-03-15
期刊：
影响因子：
64.8
通讯作者：
Ye, Yihong
中科院分区：
文献类型：
--
作者：
Li, Wei;Tu, Daqi;Ye, Yihong
在真核细胞中，许多短寿命蛋白质与Lys 48连接的泛素链结合并被蛋白酶体降解（1）。泛素化需要活化酶（E1）、结合酶（E2）和连接酶（E3）（2）。大多数泛素连接酶使用HECT（与E6相关蛋白C末端同源）或RING（真正有趣的新基因）结构域来催化多聚泛素化（3），但E3催化的机制定义不清（4）。在这里，我们使用小鼠Ube 2g 2（E2;在氨基酸水平上与人Ube 2g 2相同）和人gp 78（E3）来剖析这一过程，人gp 78是一种内质网（ER）相关的缀合系统，其对于降解错误折叠的ER蛋白是必需的（5，6）。我们通过在大肠杆菌中表达重组蛋白证明，Ube 2g 2/gp 78介导的多聚泛素化涉及在Ube 2g 2的催化半胱氨酸处预组装Lys 48连接的泛素链。Ube 2g 2锚定的泛素链的生长似乎是由两个Ube 2g 2分子之间的基于氨解的转移反应介导的，每个Ube 2g 2分子在其活性位点携带一个泛素部分。有趣的是，可以通过将预组装的遍在蛋白链从Ube 2g 2转移到底物中的赖氨酸残基来实现底物的多聚遍在蛋白化。
In eukaryotic cells, many short-lived proteins are conjugated with Lys48-linked ubiquitin chains and degraded by the proteasome(1). Ubiquitination requires an activating enzyme (E1), a conjugating enzyme (E2) and a ligase (E3)(2). Most ubiquitin ligases use either a HECT (homologous to E6-associated protein C terminus) or a RING (really interesting new gene) domain to catalyse polyubiquitination(3), but the mechanism of E3 catalysis is poorly defined(4). Here we dissect this process using mouse Ube2g2 (E2; identical at the amino acid level to human Ube2g2) and human gp78 (E3), an endoplasmic reticulum (ER)-associated conjugating system essential for the degradation of misfolded ER proteins(5,6). We demonstrate by expressing recombinant proteins in Escherichia coli that Ube2g2/gp78-mediated polyubiquitination involves preassembly of Lys48-linked ubiquitin chains at the catalytic cysteine of Ube2g2. The growth of Ube2g2-anchored ubiquitin chains seems to be mediated by an aminolysis-based transfer reaction between two Ube2g2 molecules that each carries a ubiquitin moiety in its active site. Intriguingly, polyubiquitination of a substrate can be achieved by transferring preassembled ubiquitin chains from Ube2g2 to a lysine residue in a substrate.