Synthesis of 7-aminocoumarin via Buchwald-Hartwig cross coupling for specific protein labeling in living cells
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Ting-Synthesis of 7-aminocoumarin.pdf
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Author(s)
Jin, Xin
Uttamapinant, Chayasith
Ting, Alice Y.
Date Issued
January 2011
Journal
ChemBioChem
Publisher
Wiley-VCH Verlag
Citation
Jin, Xin, Chayasith Uttamapinant, and Alice Y. Ting. “Synthesis of 7-Aminocoumarin by Buchwald-Hartwig Cross Coupling for Specific Protein Labeling in Living Cells.” ChemBioChem 12.1 (2011): 65–70.
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Author's final manuscript
Abstract
To enable minimally invasive studies of proteins in their native context, it is desirable to tag proteins with small, bright reporter groups. Recently, our lab described PRIME technology (for PRobe Incorporation Mediated by Enzymes) for such tagging[1-3]. An engineered variant of Escherichia coli lipoic acid ligase (LplA) is used to covalently attach a fluorescent substrate, such as 7-hydroxycoumarin, onto a 13-amino acid peptide recognition sequence (called LAP, for Ligase Acceptor Peptide) that is genetically fused to a protein of interest (POI) (Figure 1A). The targeting specificity is derived from the extremely high natural sequence specificity of LplA[4]. PRIME was used to label and visualize various LAP-tagged cytoskeletal and adhesion proteins in living mammalian cells. PRIME time: We report the synthesis of a pH-insensitive blue fluorophore, 7-aminocoumarin, by using palladium- catalyzed Buchwald–Hartwig cross coupling. 7-Aminocoumarin can be used to tag recombinant proteins on the cell surface and inside living cells through PRIME (probe incorporation mediated by enzymes), and unlike 7-hydroxycoumarin, can be visualized in acidic organelles such as endosomes.
MIT Department
Massachusetts Institute of Technology. Department of Chemistry
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DOI of Published Version
https://doi.org/10.1002/cbic.201000414
