Cryo-EM structure of the rhodopsin-Gαi-βγ complex reveals binding of the rhodopsin C-terminal tail to the Gβ subunit

Abstract
Data availability
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Abstract

One of the largest membrane protein families in eukaryotes are G protein-coupled receptors (GPCRs). GPCRs modulate cell physiology by activating diverse intracellular transducers, prominently heterotrimeric G proteins. The recent surge in structural data has expanded our understanding of GPCR-mediated signal transduction. However, many aspects, including the existence of transient interactions, remain elusive. We present the cryo-EM structure of the light-sensitive GPCR rhodopsin in complex with heterotrimeric Gi. Our density map reveals the receptor C-terminal tail bound to the Gβ subunit of the G protein, providing a structural foundation for the role of the C-terminal tail in GPCR signaling, and of Gβ as scaffold for recruiting Gα subunits and G protein-receptor kinases. By comparing available complexes, we found a small set of common anchoring points that are G protein-subtype specific. Taken together, our structure and analysis provide new structural basis for the molecular events of the GPCR signaling pathway.

Data availability

The cryo-EM density map of the rhodopsin-Gi complex bound to Fab16 has been deposited in the EM Data Bank (accession code EMD-4598), and the related structure coordinates have been deposited in the Protein Data Bank (accession code 6QNO). The crystal structure of Fab16 has been deposited in the Protein Data Bank (accession code 6QNK). Source data for Figure 3 is provided in Suppl. Table 3.

The following data sets were generated

1. Tsai CJ
2. Marino J
3. Adaixo RJ
4. Pamula F
5. Muehle J
6. Maeda S
7. Flock T
8. Taylor NMI
9. Mohammed I
10. Matile H
11. Dawson RJP
12. Deupi X
13. Stahlberg H
14. Schertler GFX
(2019) Rhodopsin-Gi protein complex
Electron Microscopy Data Bank, EMD-4598.

http://www.ebi.ac.uk/pdbe/entry/emdb/EMD-4598
1. Tsai CJ
2. Marino J
3. Adaixo RJ
4. Pamula F
5. Muehle J
6. Maeda S
7. Flock T
8. Taylor NMI
9. Mohammed I
10. Matile H
11. Dawson RJP
12. Deupi X
13. Stahlberg H
14. Schertler GFX
(2019) Rhodopsin-Gi protein complex
Protein Data Bank, 6QNO.

http://www.rcsb.org/structure/6QNO
1. Tsai CJ
2. Muehle J
3. Pamula F
4. Dawson RJP
5. Maeda S
6. Deupi X
7. Schertler GFX
(2019) Antibody FAB fragment targeting Gi protein heterotrimer
Protein Data Bank, 6QNK.

http://www.rcsb.org/structure/6QNK

The following previously published data sets were used

1. Tsai CJ
2. Weinert T
3. Muehle J
4. Pamula F
5. Nehme R
6. Flock T
7. Nogly P
8. Edwards PC
9. Carpenter B
10. Gruhl T
11. Ma P
12. Deupi X
13. Standfuss J
14. Tate CG
15. Schertler GFX
(2018) Crystal structure of the rhodopsin-mini-Go complex
Protein Data Bank, 6FUF.

https://www.rcsb.org/structure/6FUF
1. Lambright DG
2. Sondek J
3. Bohm A
4. Skiba NP
5. Hamm HE
6. Sigler PB
(1997) Heterotrimeric complex of a Gt-alpha/Gi-alpha chimera and the Gt-beta-gamma subunits
Protein Data Bank, 1GOT.

https://www.rcsb.org/structure/1GOT
1. Chang CC
2. Hernandez-Guzman FG
3. Luo W
4. Wang X
5. Ferrone S
6. Ghosh D
(2005) Structural basis of antigen mimicry in a clinically relevant melanoma antigen system
Protein Data Bank, 2AAB.

https://www.rcsb.org/structure/2AAB
1. Zhu Y
2. Wilson IA
(2002) Crystal structure of murine class II MHC I-Ab in complex with a human CLIP peptide
Protein Data Bank, 1MUJ.

https://www.rcsb.org/structure/1MUJ

Article and author information

Author details

Ching-Ju Tsai

Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0001-8320-5009
Jacopo Marino

Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0001-7095-0800
Ricardo Adaixo

Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Basel, Switzerland

Competing interests
No competing interests declared.
Filip Pamula

Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

Competing interests
No competing interests declared.
Jonas Muehle

Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

Competing interests
No competing interests declared.
Shoji Maeda

Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

Competing interests
No competing interests declared.
Tilman Flock

Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0003-3398-0968
Nicholas MI Taylor

Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Basel, Switzerland

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0003-0761-4921
Inayatulla Mohammed

Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Basel, Switzerland

Competing interests
No competing interests declared.
Hugues Matile

Pharma Research and Early Development, Therapeutic modalities, Roche Innovation Center Basel, Hoffmann-La Roche Ltd, Basel, Switzerland

Competing interests
No competing interests declared.
Roger JP Dawson

Pharma Research and Early Development, Therapeutic modalities, Roche Innovation Center Basel, Hoffmann-La Roche Ltd, Basel, Switzerland

Competing interests
No competing interests declared.
Xavier Deupi

Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

For correspondence
xavier.deupi@psi.ch

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0003-4572-9316
Henning Stahlberg

Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Basel, Switzerland

For correspondence
Henning.Stahlberg@unibas.ch

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0002-1185-4592
Gebhard Schertler

Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

For correspondence
gebhard.schertler@psi.ch

Competing interests
Gebhard Schertler, declares that he is a co-founder and scientific advisor of the company leadXpro AG and InterAx Biotech AG, and that he has been a member of the MAX IV Scientific Advisory Committee during the time when the research has been performed..

Funding

Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (310030_153145)

Gebhard Schertler

Swiss Nanoscience Institute (A13.12 NanoGhip)

Gebhard Schertler

National Centres of Competence in Research (TransCure)

Henning Stahlberg

Holcim Stiftung

Jacopo Marino

ETH Zurich

Tilman Flock

University of Cambridge

Tilman Flock

Roche

Shoji Maeda

Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (160805)

Xavier Deupi

Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (310030B_173335)

Gebhard Schertler

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.