Ligand modulation of sidechain dynamics in a wild-type human GPCR

Abstract
Data availability
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Abstract

GPCRs regulate all aspects of human physiology, and biophysical studies have deepened our understanding of GPCR conformational regulation by different ligands. Yet there is no experimental evidence for how sidechain dynamics control allosteric transitions between GPCR conformations. To address this deficit, we generated samples of a wild-type GPCR (A_2AR) that are deuterated apart from ¹H/¹³C NMR probes at isoleucine δ1 methyl groups, which facilitated ¹H/¹³C methyl TROSY NMR measurements with opposing ligands. Our data indicate that low [Na⁺] is required to allow large agonist-induced structural changes in A_2AR, and that patterns of sidechain dynamics substantially differ between agonist (NECA) and inverse agonist (ZM241385) bound receptors, with the inverse agonist suppressing fast ps-ns timescale motions at the G protein binding site. Our approach to GPCR NMR creates a framework for exploring how different regions of a receptor respond to different ligands or signaling proteins through modulation of fast ps-ns sidechain dynamics.

Data availability

The following data sets were generated

1. Rosenbaum et al.
(2017) NMR spectroscopy data, including wild type and mutant spectra
27261.

http://www.bmrb.wisc.edu/

The following previously published data sets were used

1. Lebon
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3. Warne
4. T.
5. Edwards
6. P.C.
7. Bennett
8. K.
9. Langmead
10. C.J.
11. Leslie
12. A.G.W.
13. Tate
14. C.G.
(2011) Thermostabilised HUMAN A2a Receptor with NECA bound
Accession number: 2YDV.

http://www.rcsb.org/pdb/explore.do?structureId=2ydv
1. Sharff
2. A.J.
3. Quiocho
4. F.A.
(1993) REFINED 1.8 ANGSTROMS STRUCTURE REVEALS THE MECHANISM OF BINDING OF A CYCLIC SUGAR, BETA-CYCLODEXTRIN, TO THE MALTODEXTRIN BINDING PROTEIN
Accession number: 1DMB.

http://www.rcsb.org/pdb/explore.do?structureId=1DMB
1. Liu
2. W.
3. Chun
4. E.
5. Thompson
6. A.A.
7. Chubukov
8. P.
9. Xu
10. F.
11. Katritch
12. V.
13. Han
14. G.W.
15. Heitman
16. L.H.
17. Ijzerman
18. A.P.
19. Cherezov
20. V.
21. Stevens
22. R.C.
23. GPCR Network
(2012) Crystal structure of the chimeric protein of A2aAR-BRIL in complex with ZM241385 at 1.8A resolution
Accession number: 4EIY.

http://www.rcsb.org/pdb/explore/explore.do?structureId=4EIY

Article and author information

Author details

Lindsay D Clark

Department of Biophysics, The University of Texas Southwestern Medical Center, Dallas, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0001-6879-6883
Igor Dikiy

Structural Biology Initiative, CUNY Advanced Science Research Center, New York, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-8155-7788
Karen Chapman

Department of Biophysics, The University of Texas Southwestern Medical Center, Dallas, United States

Competing interests
The authors declare that no competing interests exist.
Karin E J Rödström

Department of Neuroscience and Pharmacology, University of Copenhagen, Copenhagen, Denmark

Competing interests
The authors declare that no competing interests exist.
James Aramini

Structural Biology Initiative, CUNY Advanced Science Research Center, New York, United States

Competing interests
The authors declare that no competing interests exist.
Michael V LeVine

Department of Physiology and Biophysics, Weill Cornell Medical College, New York, United States

Competing interests
The authors declare that no competing interests exist.
George Khelashvili

Department of Physiology and Biophysics, Weill Cornell Medical College, New York, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0001-7235-8579
Søren G F Rasmussen

Department of Neuroscience and Pharmacology, University of Copenhagen, Copenhagen, Denmark

Competing interests
The authors declare that no competing interests exist.
Kevin H Gardner

Structural Biology Initiative, CUNY Advanced Science Research Center, New York, United States

For correspondence
Kevin.Gardner@asrc.cuny.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-8671-2556
Daniel M Rosenbaum

Department of Biophysics, The University of Texas Southwestern Medical Center, Dallas, United States

For correspondence
dan.rosenbaum@utsouthwestern.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-7174-4993

Funding

Welch Foundation (I-1770)

Daniel M Rosenbaum

National Institutes of Health (R01GM113050)

Daniel M Rosenbaum

National Institutes of Health (R01GM106239)

Kevin H Gardner

National Science Foundation (1000136529)

Lindsay D Clark

American Heart Association (16PRE27200004)

Lindsay D Clark

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

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.