title: Chemical Synthesis of K34-Ubiquitylated H2B for Nucleosome Reconstitution and Single-Particle Cryo-Electron Microscopy Structural Analysis.
abstract: 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.
authors.affiliation: 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.
authors.forename: Jiabin
authors.initials: J
authors.lastname: Li
authors.affiliation: Key Laboratory of Bioorganic Phosphorus Chemistry, Chemical Biology, Ministry of Education), Department of Chemistry and School of Life Sciences, Tsinghua University, Beijing, 100084, China.
authors.forename: Qiaoqiao
authors.initials: Q
authors.lastname: He
authors.affiliation: 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.
authors.forename: Yuntao
authors.initials: Y
authors.lastname: Liu
authors.affiliation: 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.
authors.forename: Sanling
authors.initials: S
authors.lastname: Liu
authors.affiliation: Key Laboratory of Bioorganic Phosphorus Chemistry, Chemical Biology, Ministry of Education), Department of Chemistry and School of Life Sciences, Tsinghua University, Beijing, 100084, China.
authors.forename: Shan
authors.initials: S
authors.lastname: Tang
authors.affiliation: Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.
authors.forename: Chengmin
authors.initials: C
authors.lastname: Li
authors.affiliation: 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.
authors.forename: Demeng
authors.initials: D
authors.lastname: Sun
authors.affiliation: 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.
authors.forename: Xiaorun
authors.initials: X
authors.lastname: Li
authors.affiliation: Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.
authors.forename: Min
authors.initials: M
authors.lastname: Zhou
authors.affiliation: Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, China.
authors.forename: Ping
authors.initials: P
authors.lastname: Zhu
authors.affiliation: 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.
authors.forename: Guoqiang
authors.initials: G
authors.lastname: Bi
authors.affiliation: 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.|Department of Microbiology, Immunology and Molecular Genetics and, California NanoSystems Systems, University of California Los Angeles, Los Angeles, CA, 90095, USA.
authors.forename: Zhenghong
authors.initials: Z
authors.lastname: Zhou
authors.affiliation: 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.
authors.forename: Ji-Shen
authors.initials: JS
authors.lastname: Zheng
authors.affiliation: 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.
authors.forename: Changlin
authors.initials: C
authors.lastname: Tian
mesh_terms: D020285:Cryoelectron Microscopy; D006657:Histones / Q000138:chemical synthesis* / Q000737:chemistry; D009707:Nucleosomes / Q000737:chemistry* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D054875:Ubiquitination
chemical_list: D006657:Histones; D009707:Nucleosomes
keywords: cryo-electron microscopy; histone; nucleosome; protein chemical synthesis; ubiquitylation