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Ligand-directed covalent labelling of a GPCR with a fluorescent tag
View ORCID ProfileLeigh A Stoddart, Nicholas D Kindon, Omolade Otun, Clare R. Harwood, Foteini Patera, View ORCID ProfileDmitry B Veprintsev, View ORCID ProfileJeanette Woolard, View ORCID ProfileStephen J Briddon, View ORCID ProfileHester A Franks, View ORCID ProfileStephen J Hill, View ORCID ProfileBarrie Kellam
doi: https://doi.org/10.1101/2020.04.21.053405
Leigh A Stoddart
1Cell Signalling and Pharmacology Research Group, Division of Physiology, Pharmacology and Neuroscience, School of Life Sciences, University of Nottingham, Nottingham, UK
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
Nicholas D Kindon
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
3School of Pharmacy, Biodiscovery Institute, University of Nottingham, UK
Omolade Otun
1Cell Signalling and Pharmacology Research Group, Division of Physiology, Pharmacology and Neuroscience, School of Life Sciences, University of Nottingham, Nottingham, UK
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
Clare R. Harwood
1Cell Signalling and Pharmacology Research Group, Division of Physiology, Pharmacology and Neuroscience, School of Life Sciences, University of Nottingham, Nottingham, UK
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
Foteini Patera
4Division of Cancer and Stem Cells, School of Medicine, University of Nottingham, UK
Dmitry B Veprintsev
1Cell Signalling and Pharmacology Research Group, Division of Physiology, Pharmacology and Neuroscience, School of Life Sciences, University of Nottingham, Nottingham, UK
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
Jeanette Woolard
1Cell Signalling and Pharmacology Research Group, Division of Physiology, Pharmacology and Neuroscience, School of Life Sciences, University of Nottingham, Nottingham, UK
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
Stephen J Briddon
1Cell Signalling and Pharmacology Research Group, Division of Physiology, Pharmacology and Neuroscience, School of Life Sciences, University of Nottingham, Nottingham, UK
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
Hester A Franks
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
4Division of Cancer and Stem Cells, School of Medicine, University of Nottingham, UK
Stephen J Hill
1Cell Signalling and Pharmacology Research Group, Division of Physiology, Pharmacology and Neuroscience, School of Life Sciences, University of Nottingham, Nottingham, UK
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
Barrie Kellam
2Centre of Membrane Proteins and Receptors (COMPARE), University of Birmingham and University of Nottingham, Midlands, UK
3School of Pharmacy, Biodiscovery Institute, University of Nottingham, UK

Abstract
Here, we describe rational design of a compound that covalently and selectively labels a G protein-coupled receptor (GPCR) in living cells with a fluorescent moiety. Using wild-type adenosine A2A receptor as a model system, we show fluorescent labelling without impeding access to the orthosteric binding site and demonstrate its use in endogenously expressing systems. This offers a non-invasive and selective approach to study function and localisation of native GPCRs.
Competing Interest Statement
The authors have declared no competing interest.
Copyright
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
Posted April 23, 2020.
Ligand-directed covalent labelling of a GPCR with a fluorescent tag
Leigh A Stoddart, Nicholas D Kindon, Omolade Otun, Clare R. Harwood, Foteini Patera, Dmitry B Veprintsev, Jeanette Woolard, Stephen J Briddon, Hester A Franks, Stephen J Hill, Barrie Kellam
bioRxiv 2020.04.21.053405; doi: https://doi.org/10.1101/2020.04.21.053405
Ligand-directed covalent labelling of a GPCR with a fluorescent tag
Leigh A Stoddart, Nicholas D Kindon, Omolade Otun, Clare R. Harwood, Foteini Patera, Dmitry B Veprintsev, Jeanette Woolard, Stephen J Briddon, Hester A Franks, Stephen J Hill, Barrie Kellam
bioRxiv 2020.04.21.053405; doi: https://doi.org/10.1101/2020.04.21.053405
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