组蛋白泛素化对染色体的结构和功能以及DNA相关的细胞过程具有重要的调控作用，其调控功能的异常和很多疾病的产生及发展密切相关。为理解组蛋白泛素化的功能和作用机制，特定结构的泛素修饰组蛋白是不可或缺的分子工具，如何获得位点特异性的修饰组蛋白分子是当前研究的重要挑战之一。本项目拟发展新一代辅基介导的蛋白质位点特异性泛素化合成方法，并将其应用于不同类型二泛素修饰组蛋白的制备，实现特定结构的K27或K63连接二泛素修饰H2A的化学全合成。利用泛素修饰H2A形成核小体作为分子工具，解析不同泛素修饰核小体结合53BP1的复合物结构，揭示不同位点、不同类型泛素修饰H2A调控核小体与53BP1相互作用的结构基础。研究成果将有助于理解组蛋白泛素修饰调控DNA损伤修复过程。拟发展的化学合成方法将在修饰组蛋白的制备上实现突破，在解密“组蛋白密码”研究中具有重要的应用潜力。

Histone ubiquitination plays important roles in regulating the structure and dynamics of chromatin, as well as DNA-driven cellular processes. Its dysregulation is closely related to the appearance and progression of many diseases. A key to understand the function and molecular mechanism of histone ubiquitination is the preparation of homogenous ubiquitin modified histones, which has been a challenge in this field. In this project, we plan to develop an efficient method based on auxiliary mediated ligation of peptide hydrazides for protein site-specific ubiquitination. The new method will be used in the total chemical synthesis of K27 or K63 di-ubiquitin modified H2A on Lys13/15. Then, we plan to resolve the three-dimensional structures of complexes consisting of 53BP1 bound to K27 or K63 di-ubiquitin modified nucleosome core particles (NCPs) by single particle cryo-electron microscopy, and elucidate the molecular mechanism of interaction between 53BP1 and NCP regulated by H2A ubiquitination with different linkages on different sites. The project will help to further understanding of how H2A ubiquitination regulates DNA repair. Meanwhile, the new strategy will achieve breakthroughs in the preparation of modified histones, which provides important potential applications in deciphering the “histone code”.