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- PDB-8j97: Structure of Muscarinic receptor (M2R) in complex with beta-arres... -

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Basic information

Entry
Database: PDB / ID: 8j97
TitleStructure of Muscarinic receptor (M2R) in complex with beta-arrestin1 (Local refine, cross-linked)
Components
  • Beta-arrestin-1
  • Fab30 Heavy Chain
  • Fab30 Light Chain
  • Muscarinic acetylcholine receptor M2
KeywordsSIGNALING PROTEIN/IMMUNE SYSTEM / GPCR / Arrestin / SIGNALING PROTEIN / SIGNALING PROTEIN-IMMUNE SYSTEM complex
Function / homology
Function and homology information


TGFBR3 regulates TGF-beta signaling / MAP2K and MAPK activation / Activation of SMO / Golgi Associated Vesicle Biogenesis / Lysosome Vesicle Biogenesis / Muscarinic acetylcholine receptors / G protein-coupled acetylcholine receptor activity / AP-2 adaptor complex binding / phospholipase C-activating G protein-coupled acetylcholine receptor signaling pathway / Ub-specific processing proteases ...TGFBR3 regulates TGF-beta signaling / MAP2K and MAPK activation / Activation of SMO / Golgi Associated Vesicle Biogenesis / Lysosome Vesicle Biogenesis / Muscarinic acetylcholine receptors / G protein-coupled acetylcholine receptor activity / AP-2 adaptor complex binding / phospholipase C-activating G protein-coupled acetylcholine receptor signaling pathway / Ub-specific processing proteases / clathrin coat of coated pit / clathrin heavy chain binding / cholinergic synapse / Cargo recognition for clathrin-mediated endocytosis / desensitization of G protein-coupled receptor signaling pathway / regulation of smooth muscle contraction / adenylate cyclase-inhibiting G protein-coupled acetylcholine receptor signaling pathway / Clathrin-mediated endocytosis / clathrin-dependent endocytosis / acetylcholine receptor binding / G protein-coupled receptor internalization / arrestin family protein binding / inositol hexakisphosphate binding / Thrombin signalling through proteinase activated receptors (PARs) / G alpha (s) signalling events / clathrin binding / small molecule binding / regulation of heart contraction / pseudopodium / phosphatidylinositol-3,4,5-trisphosphate binding / positive regulation of receptor internalization / negative regulation of Notch signaling pathway / G protein-coupled receptor signaling pathway, coupled to cyclic nucleotide second messenger / presynaptic modulation of chemical synaptic transmission / clathrin-coated endocytic vesicle membrane / G protein-coupled receptor binding / receptor internalization / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / response to virus / positive regulation of protein phosphorylation / G protein-coupled acetylcholine receptor signaling pathway / Cargo recognition for clathrin-mediated endocytosis / nervous system development / protein transport / presynapse / Clathrin-mediated endocytosis / cytoplasmic vesicle / ubiquitin-dependent protein catabolic process / G alpha (i) signalling events / chemical synaptic transmission / molecular adaptor activity / postsynaptic membrane / positive regulation of ERK1 and ERK2 cascade / cilium / ciliary basal body / G protein-coupled receptor signaling pathway / synapse / dendrite / nucleolus / Golgi apparatus / signal transduction / nucleus / membrane / plasma membrane / cytoplasm / cytosol
Similarity search - Function
Muscarinic acetylcholine receptor M2 / Muscarinic acetylcholine receptor family / 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 ...Muscarinic acetylcholine receptor M2 / Muscarinic acetylcholine receptor family / 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 / Serpentine type 7TM GPCR chemoreceptor Srsx / G-protein coupled receptors family 1 signature. / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / 7 transmembrane receptor (rhodopsin family) / Immunoglobulin E-set
Similarity search - Domain/homology
Muscarinic acetylcholine receptor M2 / Beta-arrestin-1
Similarity search - Component
Biological speciesBos taurus (domestic cattle)
Mus musculus (house mouse)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsMaharana, J. / Sano, F.K. / Shihoya, W. / Banerjee, R. / Nureki, O. / Shukla, A.K.
Funding support India, 4items
OrganizationGrant numberCountry
Science and Engineering Research Board (SERB)IPA/2020/000405 India
Department of Biotechnology (DBT, India)IA/S/20/1/504916 India
Science and Engineering Research Board (SERB)CRG/2022/002646 India
Science and Engineering Research Board (SERB)SPR/2020/000408 India
CitationJournal: Science / Year: 2024
Title: Molecular insights into atypical modes of β-arrestin interaction with seven transmembrane receptors.
Authors: Jagannath Maharana / Fumiya K Sano / Parishmita Sarma / Manish K Yadav / Longhan Duan / Tomasz M Stepniewski / Madhu Chaturvedi / Ashutosh Ranjan / Vinay Singh / Sayantan Saha / Gargi ...Authors: Jagannath Maharana / Fumiya K Sano / Parishmita Sarma / Manish K Yadav / Longhan Duan / Tomasz M Stepniewski / Madhu Chaturvedi / Ashutosh Ranjan / Vinay Singh / Sayantan Saha / Gargi Mahajan / Mohamed Chami / Wataru Shihoya / Jana Selent / Ka Young Chung / Ramanuj Banerjee / Osamu Nureki / Arun K Shukla /
Abstract: β-arrestins (βarrs) are multifunctional proteins involved in signaling and regulation of seven transmembrane receptors (7TMRs), and their interaction is driven primarily by agonist-induced receptor ...β-arrestins (βarrs) are multifunctional proteins involved in signaling and regulation of seven transmembrane receptors (7TMRs), and their interaction is driven primarily by agonist-induced receptor activation and phosphorylation. Here, we present seven cryo-electron microscopy structures of βarrs either in the basal state, activated by the muscarinic receptor subtype 2 (M2R) through its third intracellular loop, or activated by the βarr-biased decoy D6 receptor (D6R). Combined with biochemical, cellular, and biophysical experiments, these structural snapshots allow the visualization of atypical engagement of βarrs with 7TMRs and also reveal a structural transition in the carboxyl terminus of βarr2 from a β strand to an α helix upon activation by D6R. Our study provides previously unanticipated molecular insights into the structural and functional diversity encoded in 7TMR-βarr complexes with direct implications for exploring novel therapeutic avenues.
History
DepositionMay 2, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Dec 27, 2023Provider: repository / Type: Initial release
Revision 1.1Jan 17, 2024Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID
Revision 1.2Oct 9, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Beta-arrestin-1
H: Fab30 Heavy Chain
L: Fab30 Light Chain
V: Muscarinic acetylcholine receptor M2


Theoretical massNumber of molelcules
Total (without water)65,2464
Polymers65,2464
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Beta-arrestin-1 / Arrestin beta-1 / Arrestin-2


Mass: 39656.668 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bos taurus (domestic cattle) / Gene: ARRB1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P17870
#2: Antibody Fab30 Heavy Chain


Mass: 12861.245 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Production host: Escherichia coli (E. coli)
#3: Antibody Fab30 Light Chain


Mass: 11500.820 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Production host: Escherichia coli (E. coli)
#4: Protein/peptide Muscarinic acetylcholine receptor M2


Mass: 1226.853 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CHRM2 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P08172
Has ligand of interestY
Has protein modificationY

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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
1Muscarinic receptor (M2R) in complex with beta-arrestin1COMPLEXall0RECOMBINANT
2beta-arrestin1COMPLEX#11RECOMBINANT
3Fab30COMPLEX#2-#31RECOMBINANT
4Muscarinic receptor M2RCOMPLEX#41RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Bos taurus (domestic cattle)9913
23Mus musculus (house mouse)10090
34Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
12Spodoptera frugiperda (fall armyworm)7108
23Escherichia coli (E. coli)562
34Spodoptera frugiperda (fall armyworm)7108
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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 FIELD / Nominal defocus max: 1600 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K3 (6k x 4k) / Num. of real images: 17869
Image scansMovie frames/image: 40

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Processing

EM software
IDNameVersionCategory
4cryoSPARC3.3.1CTF correction
7Cootmodel fitting
9PHENIXmodel refinement
12cryoSPARC3.3.1classification
13cryoSPARC43D reconstruction
CTF correctionType: NONE
Particle selectionNum. of particles selected: 5235492
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 63372 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model buildingPDB-ID: 8GO8

8go8


Accession code: 8GO8 / Source name: PDB / Type: experimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0074510
ELECTRON MICROSCOPYf_angle_d0.8446150
ELECTRON MICROSCOPYf_dihedral_angle_d7.033640
ELECTRON MICROSCOPYf_chiral_restr0.048705
ELECTRON MICROSCOPYf_plane_restr0.006783

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