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- PDB-8tii: Human ACKR3 phosphorylated by GRK2 in complex with Arrestin2 in n... -

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Basic information

Entry
Database: PDB / ID: 8tii
TitleHuman ACKR3 phosphorylated by GRK2 in complex with Arrestin2 in nanodisc
Components
  • Atypical chemokine receptor 3
  • Beta-arrestin-1
  • Fab7 heavy chain
  • Fab7 light chain
  • anti-Fab nanobody
KeywordsSIGNALING PROTEIN/IMMUNE SYSTEM / complex / GPCR / arrestin / signaling / SIGNALING PROTEIN-IMMUNE SYSTEM complex
Function / homology
Function and homology information


TGFBR3 regulates TGF-beta signaling / oculomotor nerve development / MAP2K and MAPK activation / Activation of SMO / Golgi Associated Vesicle Biogenesis / Lysosome Vesicle Biogenesis / positive regulation of mesenchymal stem cell migration / C-X-C chemokine binding / AP-2 adaptor complex binding / Ub-specific processing proteases ...TGFBR3 regulates TGF-beta signaling / oculomotor nerve development / MAP2K and MAPK activation / Activation of SMO / Golgi Associated Vesicle Biogenesis / Lysosome Vesicle Biogenesis / positive regulation of mesenchymal stem cell migration / C-X-C chemokine binding / AP-2 adaptor complex binding / Ub-specific processing proteases / clathrin coat of coated pit / clathrin heavy chain binding / C-X-C chemokine receptor activity / Cargo recognition for clathrin-mediated endocytosis / desensitization of G protein-coupled receptor signaling pathway / Clathrin-mediated endocytosis / C-C chemokine receptor activity / clathrin-dependent endocytosis / negative regulation of intrinsic apoptotic signaling pathway in response to DNA damage / chemokine-mediated signaling pathway / C-C chemokine binding / G protein-coupled receptor internalization / acetylcholine receptor binding / inositol hexakisphosphate binding / Chemokine receptors bind chemokines / Thrombin signalling through proteinase activated receptors (PARs) / scavenger receptor activity / G alpha (s) signalling events / clathrin binding / small molecule binding / pseudopodium / phosphatidylinositol-3,4,5-trisphosphate binding / positive regulation of receptor internalization / negative regulation of Notch signaling pathway / vasculogenesis / coreceptor activity / clathrin-coated pit / cell chemotaxis / G protein-coupled receptor binding / calcium-mediated signaling / recycling endosome / receptor internalization / protein transport / positive regulation of protein phosphorylation / positive regulation of cytosolic calcium ion concentration / ubiquitin-dependent protein catabolic process / cytoplasmic vesicle / G alpha (i) signalling events / angiogenesis / molecular adaptor activity / early endosome / positive regulation of ERK1 and ERK2 cascade / endosome / cell adhesion / immune response / external side of plasma membrane / negative regulation of cell population proliferation / intracellular membrane-bounded organelle / cell surface / signal transduction / nucleus / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Atypical chemokine receptor 3 / : / 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 ...Atypical chemokine receptor 3 / : / 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 / 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
Beta-arrestin-1 / Atypical chemokine receptor 3
Similarity search - Component
Biological speciesBos taurus (domestic cattle)
synthetic construct (others)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å
AuthorsChen, Q. / Tesmer, J.J.G.
Funding support United States, Denmark, 12items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)CA254402 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM117372 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI161880 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA221289 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA023168 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM137505 United States
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)HL071818 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA023168 United States
Walther Cancer Foundation United States
Robertson Foundation/Cancer Research InstituteIrvington Postdoctoral Fellowship United States
VILLUM FONDEN00025326 Denmark
Ralph W. and Grace M. Showalter Research Trust4480108 United States
CitationJournal: Nature / Year: 2025
Title: Effect of phosphorylation barcodes on arrestin binding to a chemokine receptor.
Authors: Qiuyan Chen / Christopher T Schafer / Somnath Mukherjee / Kai Wang / Martin Gustavsson / James R Fuller / Katelyn Tepper / Thomas D Lamme / Yasmin Aydin / Parth Agrawal / Genki Terashi / Xin- ...Authors: Qiuyan Chen / Christopher T Schafer / Somnath Mukherjee / Kai Wang / Martin Gustavsson / James R Fuller / Katelyn Tepper / Thomas D Lamme / Yasmin Aydin / Parth Agrawal / Genki Terashi / Xin-Qiu Yao / Daisuke Kihara / Anthony A Kossiakoff / Tracy M Handel / John J G Tesmer /
Abstract: Unique phosphorylation 'barcodes' installed in different regions of an active seven-transmembrane receptor by different G-protein-coupled receptor (GPCR) kinases (GRKs) have been proposed to promote ...Unique phosphorylation 'barcodes' installed in different regions of an active seven-transmembrane receptor by different G-protein-coupled receptor (GPCR) kinases (GRKs) have been proposed to promote distinct cellular outcomes, but it is unclear whether or how arrestins differentially engage these barcodes. Here, to address this, we developed an antigen-binding fragment (Fab7) that recognizes both active arrestin2 (β-arrestin1) and arrestin3 (β-arrestin2) without interacting with bound receptor polypeptides. We used Fab7 to determine the structures of both arrestins in complex with atypical chemokine receptor 3 (ACKR3) phosphorylated in different regions of its C-terminal tail by either GRK2 or GRK5 (ref. ). The GRK2-phosphorylated ACKR3 resulted in more heterogeneous 'tail-mode' assemblies, whereas phosphorylation by GRK5 resulted in more rigid 'ACKR3-adjacent' assemblies. Unexpectedly, the finger loops of both arrestins engaged the micelle surface rather than the receptor intracellular pocket, with arrestin3 being more dynamic, partly because of its lack of a membrane-anchoring motif. Thus, both the region of the barcode and the arrestin isoform involved can alter the structure and dynamics of GPCR-arrestin complexes, providing a possible mechanistic basis for unique downstream cellular effects, such as the efficiency of chemokine scavenging and the robustness of arrestin binding in ACKR3.
History
DepositionJul 19, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 22, 2025Provider: repository / Type: Initial release
Revision 1.1Jun 18, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Beta-arrestin-1
H: Fab7 heavy chain
L: Fab7 light chain
N: anti-Fab nanobody
R: Atypical chemokine receptor 3


Theoretical massNumber of molelcules
Total (without water)154,9945
Polymers154,9945
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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


Mass: 47055.469 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bos taurus (domestic cattle) / Gene: ARRB1 / Production host: Escherichia coli (E. coli) / References: UniProt: P17870
#2: Antibody Fab7 heavy chain


Mass: 25720.758 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)
#3: Antibody Fab7 light chain


Mass: 23471.031 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)
#4: Antibody anti-Fab nanobody


Mass: 13390.644 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)
#5: Protein Atypical chemokine receptor 3 / C-X-C chemokine receptor type 7 / CXC-R7 / CXCR-7 / Chemokine orphan receptor 1 / G-protein coupled ...C-X-C chemokine receptor type 7 / CXC-R7 / CXCR-7 / Chemokine orphan receptor 1 / G-protein coupled receptor 159 / G-protein coupled receptor RDC1 homolog / RDC-1


Mass: 45356.363 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: phosphorylated by GRK2 / Source: (gene. exp.) Homo sapiens (human) / Gene: ACKR3, CMKOR1, CXCR7, GPR159, RDC1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P25106
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

ComponentName: human ACKR3 phosphorylated by GRK2 in complex with Arrestin2 in nanodisc
Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Source (natural)Organism: Bos taurus (domestic cattle)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
SpecimenConc.: 0.7 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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: 2500 nm / Nominal defocus min: 600 nm
Image recordingElectron dose: 56 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameCategory
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
CTF correctionType: NONE
3D reconstructionResolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 698683 / Symmetry type: POINT

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