Structural and functional characterization of G protein-coupled receptors with deep mutational scanning
- University of California, Los Angeles, United States
- Stanford University, United States
- MRC Laboratory of Molecular Biology, United Kingdom
Abstract
In humans, the >800 G protein-coupled receptors (GPCRs) are responsible for transducing diverse chemical stimuli to alter cell state, and are the largest class of drug targets. Their myriad structural conformations and various modes of signaling make it challenging to understand their structure and function. Here we developed a platform to characterize large libraries of GPCR variants in human cell lines with a barcoded transcriptional reporter of G-protein signal transduction. We tested 7,800 of 7,828 possible single amino acid substitutions to the beta-2 adrenergic receptor (β2AR) at four concentrations of the agonist isoproterenol. We identified residues specifically important for β2AR signaling, mutations in the human population that are potentially loss of function, and residues that modulate basal activity. Using unsupervised learning, we resolve residues critical for signaling, including all major structural motifs and molecular interfaces. We also find a previously uncharacterized structural latch spanning the first two extracellular loops that is highly conserved across Class A GPCRs and is conformationally rigid in both the inactive and active states of the receptor. More broadly, by linking deep mutational scanning with engineered transcriptional reporters, we establish a generalizable method for exploring pharmacogenomics, structure and function across broad classes of drug receptors.
Data availability
Sequencing data have been submitted to GEO and the accession code is GSE144819.
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Structural and Functional Characterization of G Protein-Coupled Receptors with Deep Mutational ScanningNCBI Gene Expression Omnibus, GSE144819.
Article and author information
Author details
A J Venkatakrishnan
Funding
National Science Foundation (1556207)
- Sriram Kosuri
National Institutes of Health (GM007185)
- Sriram Kosuri
National Institutes of Health (5T32GM008496)
- Sriram Kosuri
National Institutes of Health (DP2GM114829)
- Sriram Kosuri
Medical Research Council (MC_U105185859)
- Sriram Kosuri
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Dan Larhammar, Uppsala University, Sweden
Version history
- Received: January 5, 2020
- Accepted: October 16, 2020
- Accepted Manuscript published: October 21, 2020 (version 1)
- Version of Record published: December 1, 2020 (version 2)
Copyright
© 2020, Jones et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Structural and functional characterization of G protein-coupled receptors with deep mutational scanning
eLife 9:e54895.
https://doi.org/10.7554/eLife.54895
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