Abstract
Förster resonance energy transfer (FRET) probes are powerful tools to monitor protein–protein interactions and enzyme activities in a spatiotemporal manner in live cells. Using a combination of noncanonical amino acid (ncAA) mutagenesis and bioorthogonal labeling, we have developed intramolecular FRET probes consisting of a fluorescent protein and an organic dye within an individual protein. Herein we present a general approach to establish intramolecular FRET probes for imaging of protein activity in live cells.
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Acknowledgments
This work is supported by the Deutsche Forschungsgemeinschaft, DFG (grant No.: SPP 1623), European Research Council, ERC (ChemBioAP), and Behrens-Weise-Stifung. S.B. acknowledges support from the IMPRS-CMB. We thank Edward Lemke and Carsten Schultz for the kind gift of PylRS AF plasmid and BCN.
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Brand, S., Wu, YW. (2018). Generation of Intramolecular FRET Probes via Noncanonical Amino Acid Mutagenesis. In: Lemke, E. (eds) Noncanonical Amino Acids. Methods in Molecular Biology, vol 1728. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7574-7_21
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DOI: https://doi.org/10.1007/978-1-4939-7574-7_21
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