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- EMDB-20948: GPCR-Beta arrestin structure in lipid bilayer -

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Basic information

Entry
Database: EMDB / ID: EMD-20948
TitleGPCR-Beta arrestin structure in lipid bilayer
Map dataGPCR-Beta arrestin in lipid bilayer
Sample
  • Complex: Phosphorylated human muscarinic acetylcholine receptor M2 in complex with rat beta-arrestin1, stabilized by an antibody fragment (Fab30).
    • Complex: Phosphorylated human muscarinic acetylcholine receptor M2
    • Complex: Cysteine-free rat beta-arrestin 1 truncated at amino acid 393
    • Complex: Antibody Fragment (Fab30)
Biological speciesHomo sapiens (human) / Rattus norvegicus (Norway rat)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsStaus DP / Hu H
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood InstituteHL16037 United States
National Institutes of Health/National Institute of Neurological Disorders and StrokeNS092695 United States
CitationJournal: Nature / Year: 2020
Title: Structure of the M2 muscarinic receptor-β-arrestin complex in a lipid nanodisc.
Authors: Dean P Staus / Hongli Hu / Michael J Robertson / Alissa L W Kleinhenz / Laura M Wingler / William D Capel / Naomi R Latorraca / Robert J Lefkowitz / Georgios Skiniotis /
Abstract: After activation by an agonist, G-protein-coupled receptors (GPCRs) recruit β-arrestin, which desensitizes heterotrimeric G-protein signalling and promotes receptor endocytosis. Additionally, β- ...After activation by an agonist, G-protein-coupled receptors (GPCRs) recruit β-arrestin, which desensitizes heterotrimeric G-protein signalling and promotes receptor endocytosis. Additionally, β-arrestin directly regulates many cell signalling pathways that can induce cellular responses distinct from that of G proteins. In contrast to G proteins, for which there are many high-resolution structures in complex with GPCRs, the molecular mechanisms underlying the interaction of β-arrestin with GPCRs are much less understood. Here we present a cryo-electron microscopy structure of β-arrestin 1 (βarr1) in complex with M2 muscarinic receptor (M2R) reconstituted in lipid nanodiscs. The M2R-βarr1 complex displays a multimodal network of flexible interactions, including binding of the N domain of βarr1 to phosphorylated receptor residues and insertion of the finger loop of βarr1 into the M2R seven-transmembrane bundle, which adopts a conformation similar to that in the M2R-heterotrimeric G protein complex. Moreover, the cryo-electron microscopy map reveals that the C-edge of βarr1 engages the lipid bilayer. Through atomistic simulations and biophysical, biochemical and cellular assays, we show that the C-edge is critical for stable complex formation, βarr1 recruitment, receptor internalization, and desensitization of G-protein activation. Taken together, these data suggest that the cooperative interactions of β-arrestin with both the receptor and the phospholipid bilayer contribute to its functional versatility.
History
DepositionNov 14, 2019-
Header (metadata) releaseFeb 5, 2020-
Map releaseFeb 5, 2020-
UpdateMar 25, 2020-
Current statusMar 25, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4.2
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 4.2
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20948.png

Downloads & links

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Map

FileDownload / File: emd_20948.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationGPCR-Beta arrestin in lipid bilayer
Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 4.2 / Movie #1: 4.2
Minimum - Maximum-13.553397 - 30.015896
Average (Standard dev.)0.0070778565 (±0.83977336)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 254.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.061.061.06
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z254.400254.400254.400
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-13.55330.0160.007

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Supplemental data

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Sample components

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Entire : Phosphorylated human muscarinic acetylcholine receptor M2 in comp...

EntireName: Phosphorylated human muscarinic acetylcholine receptor M2 in complex with rat beta-arrestin1, stabilized by an antibody fragment (Fab30).
Components
  • Complex: Phosphorylated human muscarinic acetylcholine receptor M2 in complex with rat beta-arrestin1, stabilized by an antibody fragment (Fab30).
    • Complex: Phosphorylated human muscarinic acetylcholine receptor M2
    • Complex: Cysteine-free rat beta-arrestin 1 truncated at amino acid 393
    • Complex: Antibody Fragment (Fab30)

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Supramolecule #1: Phosphorylated human muscarinic acetylcholine receptor M2 in comp...

SupramoleculeName: Phosphorylated human muscarinic acetylcholine receptor M2 in complex with rat beta-arrestin1, stabilized by an antibody fragment (Fab30).
type: complex / ID: 1 / Parent: 0

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Supramolecule #2: Phosphorylated human muscarinic acetylcholine receptor M2

SupramoleculeName: Phosphorylated human muscarinic acetylcholine receptor M2
type: complex / ID: 2 / Parent: 1
Details: Phosphorylated M2R was generated by ligating a synthetic phosphopeptide derived from the vasopressin-2-receptor (V2Rpp) using the enzyme sortase.
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)

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Supramolecule #3: Cysteine-free rat beta-arrestin 1 truncated at amino acid 393

SupramoleculeName: Cysteine-free rat beta-arrestin 1 truncated at amino acid 393
type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Rattus norvegicus (Norway rat)
Recombinant expressionOrganism: Escherichia coli (E. coli)

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Supramolecule #4: Antibody Fragment (Fab30)

SupramoleculeName: Antibody Fragment (Fab30) / type: complex / ID: 4 / Parent: 1
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli)

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration2 mg/mL
BufferpH: 7.4
GridDetails: unspecified
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated magnification: 47169 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.2 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 47169
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recording#0 - Image recording ID: 1 / #0 - Film or detector model: GATAN K2 QUANTUM (4k x 4k) / #0 - Detector mode: COUNTING / #0 - Average electron dose: 50.0 e/Å2 / #1 - Image recording ID: 2 / #1 - Film or detector model: GATAN K2 QUANTUM (4k x 4k) / #1 - Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 11700000
CTF correctionSoftware - Name: Gctf (ver. 1.06)
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: RELION
Final angle assignmentType: ANGULAR RECONSTITUTION
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cisTEM / Number images used: 145618
Image recording ID1

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT

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