Li Jiabin, He Qiaoqiao, Liu Yuntao, Liu Sanling, Tang Shan, Li Chengmin, Sun Demeng, Li Xiaorun, Zhou Min, Zhu Ping, Bi Guoqiang, Zhou Zhenghong, Zheng Ji-Shen, Tian Changlin
Key Laboratory of Bioorganic Phosphorus Chemistry, Chemical Biology, Ministry of Education), Department of Chemistry and School of Life Sciences, Tsinghua University, Beijing, 100084, China.
Hefei National Laboratory of Physical Sciences at MicroScale and, School of Life Sciences, University of Science and Technology of China and, High Magnetic Field Laboratory, Chinese Academy of Sciences, Hefei, 230027, China.
Chembiochem. 2017 Jan 17;18(2):176-180. doi: 10.1002/cbic.201600551. Epub 2016 Dec 15.
Post-translational modifications (e.g., ubiquitylation) of histones play important roles in dynamic regulation of chromatin. Histone ubiquitylation has been speculated to directly influence the structure and dynamics of nucleosomes. However, structural information for ubiquitylated nucleosomes is still lacking. Here we report an alternative strategy for total chemical synthesis of homogenous histone H2B-K34-ubiquitylation (H2B-K34Ub) by using acid-cleavable auxiliary-mediated ligation of peptide hydrazides for site-specific ubiquitylation. Synthetic H2B-K34Ub was efficiently incorporated into nucleosomes and further used for single-particle cryo-electron microscopy (cryo-EM) imaging. The cryo-EM structure of the nucleosome containing H2B-K34Ub suggests that two flexible ubiquitin domains protrude between the DNA chains of the nucleosomes. The DNA chains around the H2B-K34 sites shift and provide more space for ubiquitin to protrude. These analyses indicated local and slight structural influences on the nucleosome with ubiquitylation at the H2B-K34 site.
组蛋白的翻译后修饰（如泛素化）在染色质的动态调控中发挥着重要作用。据推测，组蛋白泛素化会直接影响核小体的结构和动态变化。然而，泛素化核小体的结构信息仍然匮乏。在此，我们报告了一种通过使用可酸裂解的辅助介导肽酰肼连接进行位点特异性泛素化，来全化学合成均一的组蛋白H2B-K34-泛素化（H2B-K34Ub）的替代策略。合成的H2B-K34Ub被高效整合到核小体中，并进一步用于单颗粒冷冻电子显微镜（cryo-EM）成像。含有H2B-K34Ub的核小体的冷冻电镜结构表明，两个柔性泛素结构域在核小体的DNA链之间突出。H2B-K34位点周围的DNA链发生移位，为泛素突出提供了更多空间。这些分析表明，H2B-K34位点的泛素化对核小体有局部和轻微的结构影响。
