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- PDB-6oy9: Structure of the Rhodopsin-Transducin Complex -

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Basic information

Entry
Database: PDB / ID: 6oy9
TitleStructure of the Rhodopsin-Transducin Complex
Components
  • Gt-alpha/Gi1-alpha chimera
  • Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
  • Guanine nucleotide-binding protein G(T) subunit gamma-T1
  • Rhodopsin
KeywordsSIGNALING PROTEIN / GPCR / G protein / Complex
Function / homology
Function and homology information


sensory perception of umami taste / detection of light stimulus involved in visual perception / opsin binding / G protein-coupled receptor complex / photoreceptor connecting cilium / absorption of visible light / photoreceptor inner segment membrane / 11-cis retinal binding / negative regulation of cyclic-nucleotide phosphodiesterase activity / G protein-coupled photoreceptor activity ...sensory perception of umami taste / detection of light stimulus involved in visual perception / opsin binding / G protein-coupled receptor complex / photoreceptor connecting cilium / absorption of visible light / photoreceptor inner segment membrane / 11-cis retinal binding / negative regulation of cyclic-nucleotide phosphodiesterase activity / G protein-coupled photoreceptor activity / cellular response to light stimulus / eye photoreceptor cell development / phototransduction, visible light / arrestin family protein binding / acyl binding / photoreceptor cell maintenance / cellular response to electrical stimulus / G-protein beta/gamma-subunit complex / cellular response to catecholamine stimulus / G-protein gamma-subunit binding / outer membrane / rhodopsin mediated signaling pathway / adenylate cyclase-activating dopamine receptor signaling pathway / photoreceptor outer segment membrane / G-protein beta-subunit binding / photoreceptor disc membrane / photoreceptor outer segment / G-protein beta/gamma-subunit complex binding / heterotrimeric G-protein complex / G-protein alpha-subunit binding / response to light stimulus / phototransduction / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / cellular response to prostaglandin E stimulus / guanyl-nucleotide exchange factor activity / retina development in camera-type eye / visual perception / G protein-coupled receptor binding / photoreceptor inner segment / GDP binding / cell population proliferation / protein-chromophore linkage / cell-cell junction / GTPase activity / Golgi membrane / G protein-coupled receptor signaling pathway / GTP binding / protein phosphorylation / protein kinase binding / integral component of plasma membrane / zinc ion binding / membrane / integral component of membrane / identical protein binding / plasma membrane / metal ion binding / cytoplasm
WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase / G-protein gamma-like domain / Opsin / WD40 repeat / G-protein, beta subunit / Guanine nucleotide-binding protein, beta subunit / GPCR, rhodopsin-like, 7TM / WD40-repeat-containing domain / Rhodopsin, N-terminal ...WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase / G-protein gamma-like domain / Opsin / WD40 repeat / G-protein, beta subunit / Guanine nucleotide-binding protein, beta subunit / GPCR, rhodopsin-like, 7TM / WD40-repeat-containing domain / Rhodopsin, N-terminal / WD40 repeat, conserved site / G-protein beta WD-40 repeat / Visual pigments (opsins) retinal binding site / G-protein, gamma subunit / G-protein gamma-like domain superfamily / WD40-repeat-containing domain superfamily / G-protein alpha subunit / G-protein alpha subunit, group I / WD domain, G-beta repeat / GGL domain / Guanine nucleotide binding protein (G-protein), alpha subunit / 7 transmembrane receptor (rhodopsin family) / Amino terminal of the G-protein receptor rhodopsin / Rhodopsin / G protein-coupled receptor, rhodopsin-like / G protein alpha subunit, helical insertion / Transducin (heterotrimeric G protein), gamma chain / G Protein Gi Gamma 2 / Rhodopsin 7-helix transmembrane proteins / Rhopdopsin 7-helix transmembrane proteins / YVTN repeat-like/Quinoprotein amine dehydrogenase / 7 Propeller / Methylamine Dehydrogenase; Chain H / Few Secondary Structures / Irregular / Up-down Bundle / Mainly Beta / Mainly Alpha
Guanine nucleotide-binding protein G(t) subunit alpha-1 / Rhodopsin / Guanine nucleotide-binding protein G(T) subunit gamma-T1 / Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
Biological speciesBos taurus (cattle)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.9 Å
AuthorsGao, Y. / Hu, H. / Ramachandran, S. / Erickson, J.W. / Cerione, R.A. / Skiniotis, G.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)R01 CA201402 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01 NS092695 United States
CitationJournal: Mol. Cell / Year: 2019
Title: Structures of the Rhodopsin-Transducin Complex: Insights into G-Protein Activation.
Authors: Yang Gao / Hongli Hu / Sekar Ramachandran / Jon W Erickson / Richard A Cerione / Georgios Skiniotis /
Abstract: Rhodopsin (Rho), a prototypical G-protein-coupled receptor (GPCR) in vertebrate vision, activates the G-protein transducin (G) by catalyzing GDP-GTP exchange on its α subunit (Gα). To elucidate the ...Rhodopsin (Rho), a prototypical G-protein-coupled receptor (GPCR) in vertebrate vision, activates the G-protein transducin (G) by catalyzing GDP-GTP exchange on its α subunit (Gα). To elucidate the determinants of G coupling and activation, we obtained cryo-EM structures of a fully functional, light-activated Rho-G complex in the presence and absence of a G-protein-stabilizing nanobody. The structures illustrate how G overcomes its low basal activity by engaging activated Rho in a conformation distinct from other GPCR-G-protein complexes. Moreover, the nanobody-free structures reveal native conformations of G-protein components and capture three distinct conformers showing the Gα helical domain (αHD) contacting the Gβγ subunits. These findings uncover the molecular underpinnings of G-protein activation by visual rhodopsin and shed new light on the role played by Gβγ during receptor-catalyzed nucleotide exchange.
Validation Report
SummaryFull reportAbout validation report
History
DepositionMay 14, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 24, 2019Provider: repository / Type: Initial release
Revision 1.1Sep 4, 2019Group: Data collection / Database references / Category: citation / Item: _citation.journal_volume / _citation.page_first
Revision 1.2Nov 27, 2019Group: Data collection / Category: em_software / Item: _em_software.name
Revision 1.3Dec 4, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization

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Assembly

Deposited unit
A: Gt-alpha/Gi1-alpha chimera
B: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
G: Guanine nucleotide-binding protein G(T) subunit gamma-T1
R: Rhodopsin
hetero molecules


Theoretical massNumber of molelcules
Total (without water)127,2745
Polymers126,9894
Non-polymers2841
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area9980 Å2
ΔGint-62 kcal/mol
Surface area46010 Å2

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Components

#1: Protein Gt-alpha/Gi1-alpha chimera


Mass: 41188.895 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bos taurus (cattle) / Production host: Escherichia coli (E. coli) / References: UniProt: P04695
#2: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Transducin beta chain 1


Mass: 37430.957 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Bos taurus (cattle) / References: UniProt: P62871
#3: Protein Guanine nucleotide-binding protein G(T) subunit gamma-T1 / Transducin gamma chain


Mass: 9337.784 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Bos taurus (cattle) / References: UniProt: P02698
#4: Protein Rhodopsin /


Mass: 39031.457 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Bos taurus (cattle) / References: UniProt: P02699
#5: Chemical ChemComp-RET / RETINAL / Retinal


Mass: 284.436 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C20H28O / Feature type: SUBJECT OF INVESTIGATION

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1Rhodopsin-Transducin ComplexCOMPLEX#1-#40MULTIPLE SOURCES
2TransducinCOMPLEX#1-#31RECOMBINANT
3RhodopsinCOMPLEX#41NATURAL
Molecular weightValue: 0.1418 MDa / Experimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Bos taurus (cattle)9913
23Bos taurus (cattle)9913
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Image recordingAverage exposure time: 8 sec. / Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 12616
Image scansMovie frames/image: 40

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Processing

EM software
IDNameVersionCategory
1RELIONparticle selection
2SerialEMimage acquisition
4Gctf1.06CTF correction
12RELIONclassification
13cisTEM3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 5872324
3D reconstructionResolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 250451 / Symmetry type: POINT
Atomic model buildingProtocol: BACKBONE TRACE

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