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- EMDB-34178: Structure of beta-arrestin2 in complex with a phosphopeptide corr... -

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Basic information

Entry
Database: EMDB / ID: EMD-34178
TitleStructure of beta-arrestin2 in complex with a phosphopeptide corresponding to the human C5a anaphylatoxin chemotactic receptor 1, C5aR1
Map dataFull map with pixel size 1.2487111.
Sample
  • Complex: Peptide4 bound beta-arrestin2 in complex with Fab30
    • Complex: beta-arrestin 2
      • Protein or peptide: Beta-arrestin-2
    • Complex: Fab30
      • Protein or peptide: Fab30 Heavy Chain
      • Protein or peptide: Fab30 Light Chain
    • Complex: C5a anaphylatoxin chemotactic receptor 1
      • Protein or peptide: C5a anaphylatoxin chemotactic receptor 1
KeywordsGPCR / Arrestin / SIGNALING PROTEIN
Function / homology
Function and homology information


angiotensin receptor binding / desensitization of G protein-coupled receptor signaling pathway / G protein-coupled receptor internalization / inositol hexakisphosphate binding / phosphatidylinositol-3,4,5-trisphosphate binding / positive regulation of receptor internalization / endocytic vesicle / clathrin-coated pit / phosphatidylinositol binding / receptor internalization ...angiotensin receptor binding / desensitization of G protein-coupled receptor signaling pathway / G protein-coupled receptor internalization / inositol hexakisphosphate binding / phosphatidylinositol-3,4,5-trisphosphate binding / positive regulation of receptor internalization / endocytic vesicle / clathrin-coated pit / phosphatidylinositol binding / receptor internalization / protein transport / positive regulation of ERK1 and ERK2 cascade / signal transduction / nucleus / plasma membrane / cytoplasm
Similarity search - Function
Arrestin, conserved site / Arrestins signature. / Arrestin / Arrestin, N-terminal / Arrestin-like, N-terminal / Arrestin C-terminal-like domain / Arrestin (or S-antigen), N-terminal domain / Arrestin (or S-antigen), C-terminal domain / Arrestin (or S-antigen), C-terminal domain / Arrestin-like, C-terminal / Immunoglobulin E-set
Similarity search - Domain/homology
Biological speciesBos taurus (domestic cattle) / Mus musculus (house mouse) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.4 Å
AuthorsMaharana J / Sarma P / Yadav MK / Banerjee R / Shukla AK
Funding support India, 1 items
OrganizationGrant numberCountry
Science and Engineering Research Board (SERB)IPA/2020/000405 India
Citation
Journal: Mol Cell / Year: 2023
Title: Structural snapshots uncover a key phosphorylation motif in GPCRs driving β-arrestin activation.
Authors: Jagannath Maharana / Parishmita Sarma / Manish K Yadav / Sayantan Saha / Vinay Singh / Shirsha Saha / Mohamed Chami / Ramanuj Banerjee / Arun K Shukla /
Abstract: Agonist-induced GPCR phosphorylation is a key determinant for the binding and activation of β-arrestins (βarrs). However, it is not entirely clear how different GPCRs harboring divergent ...Agonist-induced GPCR phosphorylation is a key determinant for the binding and activation of β-arrestins (βarrs). However, it is not entirely clear how different GPCRs harboring divergent phosphorylation patterns impart converging active conformation on βarrs leading to broadly conserved functional responses such as desensitization, endocytosis, and signaling. Here, we present multiple cryo-EM structures of activated βarrs in complex with distinct phosphorylation patterns derived from the carboxyl terminus of different GPCRs. These structures help identify a P-X-P-P type phosphorylation motif in GPCRs that interacts with a spatially organized K-K-R-R-K-K sequence in the N-domain of βarrs. Sequence analysis of the human GPCRome reveals the presence of this phosphorylation pattern in a large number of receptors, and its contribution in βarr activation is demonstrated by targeted mutagenesis experiments combined with an intrabody-based conformational sensor. Taken together, our findings provide important structural insights into the ability of distinct GPCRs to activate βarrs through a significantly conserved mechanism.
#1: Journal: Mol.Cell / Year: 2023
Title: Structure of beta-arrestin in complex with a phosphopeptide
Authors: Maharana J / Sarma P / Yadav MK / Banerjee R / Shukla AK
History
DepositionAug 25, 2022-
Header (metadata) releaseMay 17, 2023-
Map releaseMay 17, 2023-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_34178.png

Downloads & links

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Map

FileDownload / File: emd_34178.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFull map with pixel size 1.2487111.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.25 Å/pix.
x 360 pix.
= 449.532 Å		1.25 Å/pix.
x 360 pix.
= 449.532 Å		1.25 Å/pix.
x 360 pix.
= 449.532 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.2487 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.12
Minimum - Maximum-0.32184315 - 0.6644659
Average (Standard dev.)-0.000015615204 (±0.013877747)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 449.532 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half map A with pixel size 1.2487111.

Fileemd_34178_half_map_1.map
AnnotationHalf map A with pixel size 1.2487111.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B with pixel size 1.2487111.

Fileemd_34178_half_map_2.map
AnnotationHalf map B with pixel size 1.2487111.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Peptide4 bound beta-arrestin2 in complex with Fab30

EntireName: Peptide4 bound beta-arrestin2 in complex with Fab30
Components
  • Complex: Peptide4 bound beta-arrestin2 in complex with Fab30
    • Complex: beta-arrestin 2
      • Protein or peptide: Beta-arrestin-2
    • Complex: Fab30
      • Protein or peptide: Fab30 Heavy Chain
      • Protein or peptide: Fab30 Light Chain
    • Complex: C5a anaphylatoxin chemotactic receptor 1
      • Protein or peptide: C5a anaphylatoxin chemotactic receptor 1

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Supramolecule #1: Peptide4 bound beta-arrestin2 in complex with Fab30

SupramoleculeName: Peptide4 bound beta-arrestin2 in complex with Fab30 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Molecular weightTheoretical: 280 KDa

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Supramolecule #2: beta-arrestin 2

SupramoleculeName: beta-arrestin 2 / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Bos taurus (domestic cattle)

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Supramolecule #3: Fab30

SupramoleculeName: Fab30 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2-#3
Source (natural)Organism: Mus musculus (house mouse)

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Supramolecule #4: C5a anaphylatoxin chemotactic receptor 1

SupramoleculeName: C5a anaphylatoxin chemotactic receptor 1 / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #4
Source (natural)Organism: Homo sapiens (human) / Synthetically produced: Yes

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Macromolecule #1: Beta-arrestin-2

MacromoleculeName: Beta-arrestin-2 / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (domestic cattle)
Molecular weightTheoretical: 47.217676 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MGEKPGTRVF KKSSPNGKLT VYLGKRDFVD HLDKVDPVDG VVLVDPDYLK DRKVFVTLTV AFRYGREDCD VLGLSFRKDL FIANYQAFP PTPNPPRPPT RLQERLLRKL GQHAHPFFFT IPQNLPSSVT LQPGPEDTGK ALGVDFEIRA FVAKSLEEKS H KRNSVRLV ...String:
MGEKPGTRVF KKSSPNGKLT VYLGKRDFVD HLDKVDPVDG VVLVDPDYLK DRKVFVTLTV AFRYGREDCD VLGLSFRKDL FIANYQAFP PTPNPPRPPT RLQERLLRKL GQHAHPFFFT IPQNLPSSVT LQPGPEDTGK ALGVDFEIRA FVAKSLEEKS H KRNSVRLV IRKVQFAPEK PGPQPSAETT RHFLMSDRSL HLEASLDKEL YYHGEPLNVN VHVTNNSTKT VKKIKVSVRQ YA DIVLFST AQYKVPVAQV EQDDQVSPSS TFSKVYTITP FLANNREKRG LALDGKLKHE DTNLASSTIV KEGANKEVLG ILV SYRVKV KLVVSRGGDV SVELPFVLMH PKPHDHIALP RPQSAATHPP TLLPSAVPET DAPVDTNLIE FETNYATDDD IVFE DFARL RLKGLKDEDY DDQFC

UniProtKB: Beta-arrestin-2

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Macromolecule #2: Fab30 Heavy Chain

MacromoleculeName: Fab30 Heavy Chain / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 25.512354 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: EISEVQLVES GGGLVQPGGS LRLSCAASGF NVYSSSIHWV RQAPGKGLEW VASISSYYGY TYYADSVKGR FTISADTSKN TAYLQMNSL RAEDTAVYYC ARSRQFWYSG LDYWGQGTLV TVSSASTKGP SVFPLAPSSK STSGGTAALG CLVKDYFPEP V TVSWNSGA ...String:
EISEVQLVES GGGLVQPGGS LRLSCAASGF NVYSSSIHWV RQAPGKGLEW VASISSYYGY TYYADSVKGR FTISADTSKN TAYLQMNSL RAEDTAVYYC ARSRQFWYSG LDYWGQGTLV TVSSASTKGP SVFPLAPSSK STSGGTAALG CLVKDYFPEP V TVSWNSGA LTSGVHTFPA VLQSSGLYSL SSVVTVPSSS LGTQTYICNV NHKPSNTKVD KKVEPKSCDK THHHHHHHH

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Macromolecule #3: Fab30 Light Chain

MacromoleculeName: Fab30 Light Chain / type: protein_or_peptide / ID: 3 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 23.435064 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: SDIQMTQSPS SLSASVGDRV TITCRASQSV SSAVAWYQQK PGKAPKLLIY SASSLYSGVP SRFSGSRSGT DFTLTISSLQ PEDFATYYC QQYKYVPVTF GQGTKVEIKR TVAAPSVFIF PPSDSQLKSG TASVVCLLNN FYPREAKVQW KVDNALQSGN S QESVTEQD ...String:
SDIQMTQSPS SLSASVGDRV TITCRASQSV SSAVAWYQQK PGKAPKLLIY SASSLYSGVP SRFSGSRSGT DFTLTISSLQ PEDFATYYC QQYKYVPVTF GQGTKVEIKR TVAAPSVFIF PPSDSQLKSG TASVVCLLNN FYPREAKVQW KVDNALQSGN S QESVTEQD SKDSTYSLSS TLTLSKADYE KHKVYACEVT HQGLSSPVTK SFNRGEC

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Macromolecule #4: C5a anaphylatoxin chemotactic receptor 1

MacromoleculeName: C5a anaphylatoxin chemotactic receptor 1 / type: protein_or_peptide / ID: 4 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 2.69834 KDa
SequenceString:
E(SEP)K(SEP)F(TPO)R(SEP)(TPO)V D(TPO)MAQKTQAV

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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

BufferpH: 7.4
Component:
ConcentrationFormulaName
20.0 mMC8H18N2O4SHEPES
300.0 mMNaClSodium chloride
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 283.15 K / Instrument: LEICA EM GP / Details: Blotted for 3 seconds before plunging..

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Electron microscopy

MicroscopeTFS GLACIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Detector mode: COUNTING / Number real images: 8614 / Average electron dose: 51.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 46000
Sample stageCooling holder cryogen: NITROGEN

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Image processing

Particle selectionNumber selected: 4012616
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

8goc

Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.1) / Number images used: 38206
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final 3D classificationSoftware - Name: cryoSPARC (ver. 3.3.1)

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Atomic model buiding 1

Initial modelPDB ID:

8goc


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-8goo:
Structure of beta-arrestin2 in complex with a phosphopeptide corresponding to the human C5a anaphylatoxin chemotactic receptor 1, C5aR1

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