Genetically encoding ε-N-methacryllysine into proteins in live cells
Lysine acylation is a ubiquitous post-translational modification (PTM) that plays pivotal roles in various cellular processes, such as transcription, metabolism, protein localization and folding. Thousands of lysine acylation sites have been identified based on advances in antibody enrichment strategies, highly sensitive analysis by mass spectrometry (MS), and bioinformatics. However, only 27 lysine methacrylation (Kmea) sites have been identified exclusively in histone proteins. It is hard to separate, purify and differentiate the Kmea modification from its structural isomer lysine crotonylation (Kcr) using general biochemical approaches. Here, we identify Kmea sites on a non-histone protein, Cyclophillin A (CypA). To investigate the functions of Kmea in CypA, we develop a general genetic code expansion approach to incorporate a non-canonical amino acid (ncAA) ε-N-Methacryllysine (MeaK) into target proteins and identify interacting proteins of methacrylated CypA using affinity-purification MS. We find that Kmea at CypA site 125 regulates cellular redox homeostasis, and HDAC1 is the regulator of Kmea on CypA. Moreover, we discover that genetically encode Kmea can be further methylated to ε-N-methyl-ε-N-methacrylation (Kmemea) in live cells.
Authors
- Haifan Wu (ORCID: https://orcid.org/0000-0002-2050-9950)
- Emmanuel Gboyega Ajiboye
- Yaxin Fu (ORCID: https://orcid.org/0009-0008-2959-4216)
- Anna Sablina (ORCID: https://orcid.org/0000-0001-9526-4014)
- Ying Yuan (ORCID: https://orcid.org/0000-0002-3922-9553)
- Xintong Zhou
- Tianyi Zhu (ORCID: https://orcid.org/0000-0001-9291-8975)
- Ming Wu (ORCID: https://orcid.org/0000-0002-1009-5387)
- Bing Yang (ORCID: https://orcid.org/0000-0001-6739-2932)
- Hai Song (ORCID: https://orcid.org/0000-0003-3228-6616)
- Heyu Li
- Bo Jin (ORCID: https://orcid.org/0000-0001-9386-8471)
- Dandan Liu (ORCID: https://orcid.org/0000-0002-2090-9217)
- Long Zhang (ORCID: https://orcid.org/0000-0001-8139-0474)
- Shiyi Chen (ORCID: https://orcid.org/0000-0002-3955-0101)
- Han Huang (ORCID: https://orcid.org/0000-0003-3353-2970)
- Mengdi Zhang (ORCID: https://orcid.org/0000-0001-5752-9464)
- Ke Sun (ORCID: https://orcid.org/0009-0004-4700-9077)
- Ting Wu (ORCID: https://orcid.org/0000-0002-0434-3700)
- Peilong Lu (ORCID: https://orcid.org/0000-0001-5894-9268)
- Xiaobo Wan (ORCID: https://orcid.org/0000-0001-5740-512X)
- Chao Tong
- Jia Wen Wang
Institutions
- Capital Medical University (CN)
- Westlake University (CN)
- Wichita State University (US)
- VIB-KU Leuven Center for Cancer Biology (BE)
- Zhejiang Cancer Hospital (CN)
- Beijing Computational Science Research Center (CN)
- Second Affiliated Hospital of Zhejiang University (CN)
- Zhejiang University (CN)
Publication Details
- Journal
- Nature Communications
- Published
- 2025-03-17
- DOI
- https://doi.org/10.1038/s41467-025-57969-2
- Citations
- 2
- Primary Topic
- RNA Interference and Gene Delivery
- Type
- article
- Field-Weighted Citation Impact
- 1.61