Journal
CHEM
Volume 5, Issue 11, Pages 2955-2968Publisher
CELL PRESS
DOI: 10.1016/j.chempr.2019.08.020
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Funding
- National Science Foundation of China (NSFC) [21602235, 21778062]
- Chinese Academy of Science
- Natural Science Foundation of Shanghai [16ZR1442800]
- Shanghai Rising-Star program
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The genetically encoded non-selective photo-crosslinkers suffer from unpredictable crosslinking sites and false positive interacting proteins, thus, the identification of protein interactions and mapping accurate interaction interfaces remain largely elusive in living systems. Here, we report genetically encoded, residue-selective protein photo-crosslinker enabling covalent crosslinking of the interacting proteins with proximal lysine, upon UV light activation in vitro and in living cells. Using this photo-crosslinker, we demonstrate a crosslinking strategy to capture protein-protein interactions with residue selectivity in a temporal-resolved manner. In addition, this crosslinking strategy produces the lysine-selective crosslinking site and predictable crosslinked peptide, which serve as the direct evidence for protein-protein interactions and facilitate mass spectrometry analysis. More importantly, this photo-crosslinker enables the capture of elusive enzyme-substrate interaction with directly interacting lysine and validation of acetylation site of the substrate. This strategy represents higher spatiotemporal resolution, accuracy, and reliability for investigating dynamic, transient, and weak protein-protein interactions in living systems.
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