Huang, Yang, Wu, Chengyang, Lu, Anjing, Wang, Jingzhe, Liang, Jian, Sun, Han, Yang, Liqing, Duan, Shixiang, Berezin, Andrey A., Wu, Chuanliu, Zhang, Bo, Wu, Yi-Lin  ORCID: https://orcid.org/0000-0003-0253-1625 and Tsai, Yu-Hsuan
2025.
A single bioorthogonal reaction for multiplex cell surface protein labeling.
Journal of the American Chemical Society
147
(2)
, 1612–1623.
10.1021/jacs.4c11701
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Abstract
Small-molecule fluorophores are invaluable tools for fluorescence imaging. However, means for their covalent conjugation to the target proteins limit applications in multicolor imaging. Here, we identify 2-[(alkylthio)(aryl)methylene]malononitrile (TAMM) molecules reacting with 1,2-aminothiol at a labeling rate over 104 M–1 s–1 through detailed mechanistic investigation. The fast TAMM molecules and mild reaction conditions enable site-specific labeling of surface proteins in not only cell lines but also primary neurons and living mice. The combination of genetic code expansion and sequence-specific proteolytic cleavage enables selective modification of three different cell surface proteins through iterative TAMM condensation. TAMM condensation is also compatible with Cu-catalyzed azide–alkyne cycloaddition and tetrazine ligation for four-color fluorescent labeling, reaching the maximum available colors of conventional confocal microscopes. Thus, bioconjugation chemistry is no longer the limiting factor for multiplex cell surface protein imaging.
| Item Type: | Article |
|---|---|
| Date Type: | Publication |
| Status: | Published |
| Schools: | Professional Services > Advanced Research Computing @ Cardiff (ARCCA) Schools > Chemistry |
| Publisher: | American Chemical Society |
| ISSN: | 0002-7863 |
| Funders: | National Natural Science Foundation of China |
| Date of First Compliant Deposit: | 6 January 2025 |
| Date of Acceptance: | 18 December 2024 |
| Last Modified: | 01 Jul 2025 16:01 |
| URI: | https://orca.cardiff.ac.uk/id/eprint/175003 |
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