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- PDB-7uxf: Cryogenic electron microscopy 3D map of F-actin -

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Basic information

Entry
Database: PDB / ID: 7uxf
TitleCryogenic electron microscopy 3D map of F-actin
ComponentsActin, alpha skeletal muscle
KeywordsCELL ADHESION / F-actin / cell-cell junction
Function / homology
Function and homology information


cytoskeletal motor activator activity / tropomyosin binding / myosin heavy chain binding / mesenchyme migration / troponin I binding / actin filament bundle / filamentous actin / actin filament bundle assembly / skeletal muscle thin filament assembly / striated muscle thin filament ...cytoskeletal motor activator activity / tropomyosin binding / myosin heavy chain binding / mesenchyme migration / troponin I binding / actin filament bundle / filamentous actin / actin filament bundle assembly / skeletal muscle thin filament assembly / striated muscle thin filament / skeletal muscle myofibril / actin monomer binding / skeletal muscle fiber development / stress fiber / titin binding / actin filament polymerization / filopodium / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / calcium-dependent protein binding / lamellipodium / cell body / hydrolase activity / protein domain specific binding / calcium ion binding / positive regulation of gene expression / magnesium ion binding / ATP binding / identical protein binding / cytoplasm
Similarity search - Function
Actins signature 1. / Actin, conserved site / Actins signature 2. / Actin/actin-like conserved site / Actins and actin-related proteins signature. / Actin / Actin family / Actin / ATPase, nucleotide binding domain
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / Actin, alpha skeletal muscle
Similarity search - Component
Biological speciesOryctolagus cuniculus (rabbit)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsRangarajan, E.S. / Smith, E.W. / Izard, T.
Funding support United States, 1items
OrganizationGrant numberCountry
National Science Foundation (NSF, United States) United States
CitationJournal: Commun Biol / Year: 2023
Title: Distinct inter-domain interactions of dimeric versus monomeric α-catenin link cell junctions to filaments.
Authors: Erumbi S Rangarajan / Emmanuel W Smith / Tina Izard /
Abstract: Attachment between cells is crucial for almost all aspects of the life of cells. These inter-cell adhesions are mediated by the binding of transmembrane cadherin receptors of one cell to cadherins of ...Attachment between cells is crucial for almost all aspects of the life of cells. These inter-cell adhesions are mediated by the binding of transmembrane cadherin receptors of one cell to cadherins of a neighboring cell. Inside the cell, cadherin binds β-catenin, which interacts with α-catenin. The transitioning of cells between migration and adhesion is modulated by α-catenin, which links cell junctions and the plasma membrane to the actin cytoskeleton. At cell junctions, a single β-catenin/α-catenin heterodimer slips along filamentous actin in the direction of cytoskeletal tension which unfolds clustered heterodimers to form catch bonds with F-actin. Outside cell junctions, α-catenin dimerizes and links the plasma membrane to F-actin. Under cytoskeletal tension, α-catenin unfolds and forms an asymmetric catch bond with F-actin. To understand the mechanism of this important α-catenin function, we determined the 2.7 Å cryogenic electron microscopy (cryoEM) structures of filamentous actin alone and bound to human dimeric α-catenin. Our structures provide mechanistic insights into the role of the α-catenin interdomain interactions in directing α-catenin function and suggest a bivalent mechanism. Further, our cryoEM structure of human monomeric α-catenin provides mechanistic insights into α-catenin autoinhibition. Collectively, our structures capture the initial α-catenin interaction with F-actin before the sensing of force, which is a crucial event in cell adhesion and human disease.
History
DepositionMay 5, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 8, 2023Provider: repository / Type: Initial release
Revision 1.1Mar 29, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _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

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Actin, alpha skeletal muscle
B: Actin, alpha skeletal muscle
C: Actin, alpha skeletal muscle
D: Actin, alpha skeletal muscle
E: Actin, alpha skeletal muscle
F: Actin, alpha skeletal muscle
hetero molecules


Theoretical massNumber of molelcules
Total (without water)255,36918
Polymers252,6606
Non-polymers2,70912
Water1,08160
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1chain "C"
d_2ens_1chain "B"
d_3ens_1chain "D"
d_4ens_1chain "A"
d_5ens_1chain "E"
d_6ens_1chain "F"

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg label comp-IDEnd label comp-IDLabel asym-IDLabel seq-ID
d_11ens_1THRCYSE1 - 370
d_12ens_1ADPADPF
d_21ens_1THRCYSC1 - 370
d_22ens_1ADPADPD
d_31ens_1THRCYSG1 - 370
d_32ens_1ADPADPH
d_41ens_1THRCYSA1 - 370
d_42ens_1ADPADPB
d_51ens_1THRCYSI1 - 370
d_52ens_1ADPADPJ
d_61ens_1THRCYSK1 - 370
d_62ens_1ADPADPL

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Components

#1: Protein
Actin, alpha skeletal muscle / / Alpha-actin-1


Mass: 42109.973 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Oryctolagus cuniculus (rabbit) / Gene: ACTA1, ACTA / Production host: Escherichia coli (E. coli) / References: UniProt: P68135
#2: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE / Adenosine diphosphate


Mass: 427.201 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Comment: ADP, energy-carrying molecule*YM
#3: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Mg
#4: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 60 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: FILAMENT / 3D reconstruction method: helical reconstruction

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Sample preparation

ComponentName: dimeric alpha-catenin (residues 22-906) complexed with F-actin
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Oryctolagus cuniculus (rabbit)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
1150 mMSodium ChlorideNaClSodium chloride1
220 mMTrisTris1
31 mMDithiothreitolDTT1
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 65 % / Chamber temperature: 294.15 K

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Electron microscopy imaging

MicroscopyModel: JEOL CRYO ARM 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 60000 X / Nominal defocus max: 3000 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: JEOL CRYOSPECPORTER
Image recordingAverage exposure time: 0.1 sec. / Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)
EM imaging opticsEnergyfilter name: In-column Omega Filter / Phase plate: OTHER

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.18.2_3874refinement
PHENIX1.18.2_3874refinement
EM software
IDNameCategory
2SerialEMimage acquisition
4cryoSPARCCTF correction
7UCSF Chimeramodel fitting
9Cootmodel refinement
10PHENIXmodel refinement
11cryoSPARCinitial Euler assignment
12cryoSPARCfinal Euler assignment
14cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Helical symmertyAngular rotation/subunit: -166.7 ° / Axial rise/subunit: 27.4 Å / Axial symmetry: C2
3D reconstructionResolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 963102 / Symmetry type: HELICAL
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingPDB-ID: 7UTJ

7utj


Accession code: 7UTJ / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 42.36 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00717874
ELECTRON MICROSCOPYf_angle_d0.72124258
ELECTRON MICROSCOPYf_dihedral_angle_d24.5252490
ELECTRON MICROSCOPYf_chiral_restr0.052700
ELECTRON MICROSCOPYf_plane_restr0.0053096
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDRefine-IDTypeRms dev position (Å)
ens_1d_2EELECTRON MICROSCOPYNCS constraints0.000706198487741
ens_1d_3EELECTRON MICROSCOPYNCS constraints0.000705670835478
ens_1d_4EELECTRON MICROSCOPYNCS constraints0.000710930675981
ens_1d_5EELECTRON MICROSCOPYNCS constraints0.0675011844545
ens_1d_6EELECTRON MICROSCOPYNCS constraints0.0894876828986

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