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Optimizing the Genetic Incorporation of Chemical Probes into GPCRs for Photo-crosslinking Mapping and Bioorthogonal Chemistry in Live Mammalian Cells
 

Optimizing the Genetic Incorporation of Chemical Probes into GPCRs for Photo-crosslinking Mapping and Bioorthogonal Chemistry in Live Mammalian Cells

Article DOI: 10.3791/57069-v • 14:02 min • April 9th, 2018
April 9th, 2018 •
*1, *1, *1, 1
1Institute of Biochemistry, Faculty of Life Sciences, University of Leipzig
* These authors contributed equally

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A facile fluorescence assay is presented to evaluate the efficiency of amino-acyl-tRNA-synthetase/tRNA pairs incorporating non-canonical amino-acids (ncAAs) into proteins expressed in mammalian cells. The application of ncAAs to study G-protein coupled receptors (GPCRs) is described, including photo-crosslinking mapping of binding sites and bioorthogonal GPCR labeling on live cells.

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Genetic Incorporation Chemical Probes GPCRs Photo-crosslinking Mapping Bioorthogonal Chemistry Live Mammalian Cells Non-canonical Amino Acids Protein-protein Interaction Surfaces Amber Codon Suppression Fluorescence Assay Ligand Binding Sites Intake Receptor Bioorthogonal Labeling Site-specific Labeling Catalyst-free Labeling Small Fluorophores High-end Imaging Applications
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