Engineering Research Center of Cell and Therapeutic Antibody, Ministry of Education, and School of Pharmacy, Shanghai Jiao Tong University, Shanghai, China (B.Z., B.J., B.W.); Department of Pathophysiology, School of Medicine, Jinan University, Guangzhou, China (Y.W.); Laboratory of Protein Dynamics and Signaling, Center for Cancer Research, National Cancer Institute, Frederick, Maryland (Y.C.T., A.M.W.); and Department of Chemistry, Center for Diagnostics and Therapeutics, Georgia State University, Atlanta, Georgia (Y.W., H.Z., T.C., J.Y.)
Engineering Research Center of Cell and Therapeutic Antibody, Ministry of Education, and School of Pharmacy, Shanghai Jiao Tong University, Shanghai, China (B.Z., B.J., B.W.); Department of Pathophysiology, School of Medicine, Jinan University, Guangzhou, China (Y.W.); Laboratory of Protein Dynamics and Signaling, Center for Cancer Research, National Cancer Institute, Frederick, Maryland (Y.C.T., A.M.W.); and Department of Chemistry, Center for Diagnostics and Therapeutics, Georgia State University, Atlanta, Georgia (Y.W., H.Z., T.C., J.Y.).
Pharmacol Rev. 2020 Apr;72(2):380-413. doi: 10.1124/pr.118.015651.
Ubiquitin (UB) transfer cascades consisting of E1, E2, and E3 enzymes constitute a complex network that regulates a myriad of biologic processes by modifying protein substrates. Deubiquitinating enzymes (DUBs) reverse UB modifications or trim UB chains of diverse linkages. Additionally, many cellular proteins carry UB-binding domains (UBDs) that translate the signals encoded in UB chains to target proteins for degradation by proteasomes or in autophagosomes, as well as affect nonproteolytic outcomes such as kinase activation, DNA repair, and transcriptional regulation. Dysregulation of the UB transfer pathways and malfunctions of DUBs and UBDs play causative roles in the development of many diseases. A greater understanding of the mechanism of UB chain assembly and the signals encoded in UB chains should aid in our understanding of disease pathogenesis and guide the development of novel therapeutics. The recent flourish of protein-engineering approaches such as unnatural amino acid incorporation, protein semisynthesis by expressed protein ligation, and high throughput selection by phage and yeast cell surface display has generated designer proteins as powerful tools to interrogate cell signaling mediated by protein ubiquitination. In this study, we highlight recent achievements of protein engineering on mapping, probing, and manipulating UB transfer in the cell. SIGNIFICANCE STATEMENT: The post-translational modification of proteins with ubiquitin alters the fate and function of proteins in diverse ways. Protein engineering is fundamentally transforming research in this area, providing new mechanistic insights and allowing for the exploration of concepts that can potentially be applied to therapeutic intervention.
泛素 (UB) 转移级联反应由 E1、E2 和 E3 酶组成，构成了一个复杂的网络，通过修饰蛋白质底物来调节无数的生物过程。去泛素化酶 (DUB) 逆转 UB 修饰或修剪具有不同连接的 UB 链。此外，许多细胞蛋白携带 UB 结合结构域 (UBD)，将 UB 链中编码的信号翻译成靶蛋白，通过蛋白酶体或自噬体进行降解，以及影响非蛋白水解的结果，如激酶激活、DNA 修复和转录调节。UB 转移途径的失调和 DUB 和 UBD 的功能障碍在许多疾病的发展中起因果作用。对 UB 链组装机制和 UB 链中编码的信号的更好理解应该有助于我们了解疾病的发病机制，并指导新型治疗方法的开发。最近，蛋白质工程方法的蓬勃发展，如非天然氨基酸掺入、通过表达蛋白连接的蛋白质半合成以及噬菌体和酵母细胞表面展示的高通量选择，产生了设计蛋白，作为研究蛋白质泛素化介导的细胞信号的有力工具。在本研究中，我们强调了蛋白质工程在细胞中对 UB 转移进行映射、探测和操作方面的最新成就。意义陈述：蛋白质与泛素的翻译后修饰以多种方式改变蛋白质的命运和功能。蛋白质工程从根本上改变了该领域的研究，提供了新的机制见解，并允许探索可能应用于治疗干预的概念。
