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An E1-Catalyzed Chemoenzymatic Strategy to Isopeptide-N-Ethylated Deubiquitylase-Resistant Ubiquitin Probes |
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| Authors: | Qingyun Zheng Tian Wang Guo-Chao Chu Chong Zuo Rui Zhao Xin Sui Linzhi Ye Yuanyuan Yu Jingnan Chen Xiangwei Wu Wenhao Zhang Prof Haiteng Deng Prof Jing Shi Dr Man Pan Prof Yi-Ming Li Prof Lei Liu |
| Institution: | 1. Tsinghua-Peking Center for Life Sciences, Ministry of Education Key Laboratory of Bioorganic Phosphorus Chemistry and Chemical Biology, Center for Synthetic and Systems Biology, Department of Chemistry, Tsinghua University, Beijing, 100084 China
These authors contributed equally to this work.;2. School of Food and Biological Engineering, Hefei University of Technology, Hefei, 230009 China Department of Chemistry, University of Science and Technology of China, Hefei, 230026 China These authors contributed equally to this work.;3. Tsinghua-Peking Center for Life Sciences, Ministry of Education Key Laboratory of Bioorganic Phosphorus Chemistry and Chemical Biology, Center for Synthetic and Systems Biology, Department of Chemistry, Tsinghua University, Beijing, 100084 China;4. School of Food and Biological Engineering, Hefei University of Technology, Hefei, 230009 China Department of Chemistry, University of Science and Technology of China, Hefei, 230026 China;5. School of Food and Biological Engineering, Hefei University of Technology, Hefei, 230009 China;6. MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing, 100084 China;7. Department of Chemistry, University of Science and Technology of China, Hefei, 230026 China;8. Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, IL, 60637 USA |
| Abstract: | Triazole-based deubiquitylase (DUB)-resistant ubiquitin (Ub) probes have recently emerged as effective tools for the discovery of Ub chain-specific interactors in proteomic studies, but their structural diversity is limited. A new family of DUB-resistant Ub probes is reported based on isopeptide-N-ethylated dimeric or polymeric Ub chains, which can be efficiently prepared by a one-pot, ubiquitin-activating enzyme (E1)-catalyzed condensation reaction of recombinant Ub precursors to give various homotypic and even branched Ub probes at multi-milligram scale. Proteomic studies using label-free quantitative (LFQ) MS indicated that the isopeptide-N-ethylated Ub probes may complement the triazole-based probes in the study of Ub interactome. Our study highlights the utility of modern protein synthetic chemistry to develop structurally and new families of tool molecules needed for proteomic studies. |
| Keywords: | interactomes probes proteomics ubiquitin |