Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL, USA.
Tsinghua-Peking Center for Life Sciences, Department of Chemistry, Tsinghua University, Beijing, China.
Nat Chem Biol. 2021 Aug;17(8):896-905. doi: 10.1038/s41589-021-00823-5. Epub 2021 Jul 8.
Protein ubiquitination shows remarkable topological and functional diversity through the polymerization of ubiquitin via different linkages. Deciphering the cellular ubiquitin code is of central importance to understand the physiology of the cell. However, our understanding of its function is rather limited due to the lack of specific binders as tools to detect K29-linked polyubiquitin. In this study, we screened and characterized a synthetic antigen-binding fragment, termed sAB-K29, that can specifically recognize K29-linked polyubiquitin using chemically synthesized K29-linked diubiquitin. We further determined the crystal structure of this fragment bound to the K29-linked diubiquitin, which revealed the molecular basis of specificity. Using sAB-K29 as a tool, we uncovered that K29-linked ubiquitination is involved in different kinds of cellular proteotoxic stress response as well as cell cycle regulation. In particular, we showed that K29-linked ubiquitination is enriched in the midbody and downregulation of the K29-linked ubiquitination signal arrests cells in G1/S phase.
蛋白质泛素化通过不同连接的泛素聚合显示出显著的拓扑和功能多样性。解析细胞内的泛素密码对于理解细胞生理学至关重要。然而，由于缺乏特异性结合物作为检测 K29 连接多泛素的工具，我们对其功能的理解相当有限。在这项研究中，我们筛选和表征了一种合成抗原结合片段，称为 sAB-K29，它可以使用化学合成的 K29 连接二泛素来特异性识别 K29 连接多泛素。我们进一步确定了该片段与 K29 连接二泛素结合的晶体结构，揭示了特异性的分子基础。使用 sAB-K29 作为工具，我们发现 K29 连接的泛素化参与了不同类型的细胞蛋白毒性应激反应以及细胞周期调控。特别是，我们表明 K29 连接的泛素化在中体中富集，并且下调 K29 连接的泛素化信号会导致细胞停滞在 G1/S 期。
