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- PDB-9az6: F-actin-Talin(R13-DD) complex -

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Basic information

Entry
Database: PDB / ID: 9az6
TitleF-actin-Talin(R13-DD) complex
Components
  • Actin, alpha skeletal muscle
  • Talin-1
KeywordsCELL ADHESION / actin / talin / focal adhesion / tension sensor / ABS3 / single particle
Function / homology
Function and homology information


GRB2:SOS provides linkage to MAPK signaling for Integrins / Integrin signaling / p130Cas linkage to MAPK signaling for integrins / SEMA3A-Plexin repulsion signaling by inhibiting Integrin adhesion / MAP2K and MAPK activation / LIM domain binding / Smooth Muscle Contraction / Platelet degranulation / cortical microtubule organization / vinculin binding ...GRB2:SOS provides linkage to MAPK signaling for Integrins / Integrin signaling / p130Cas linkage to MAPK signaling for integrins / SEMA3A-Plexin repulsion signaling by inhibiting Integrin adhesion / MAP2K and MAPK activation / LIM domain binding / Smooth Muscle Contraction / Platelet degranulation / cortical microtubule organization / vinculin binding / integrin activation / cell-substrate junction assembly / cortical actin cytoskeleton organization / cytoskeletal motor activator activity / phosphatidylserine binding / myosin heavy chain binding / tropomyosin binding / troponin I binding / filamentous actin / mesenchyme migration / actin filament bundle / actin filament bundle assembly / skeletal muscle myofibril / striated muscle thin filament / skeletal muscle thin filament assembly / actin monomer binding / stress fiber / skeletal muscle fiber development / ruffle / titin binding / actin filament polymerization / phosphatidylinositol binding / integrin-mediated signaling pathway / filopodium / adherens junction / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / structural constituent of cytoskeleton / ruffle membrane / calcium-dependent protein binding / actin filament binding / integrin binding / lamellipodium / cell body / cytoskeleton / hydrolase activity / cell adhesion / protein domain specific binding / focal adhesion / calcium ion binding / positive regulation of gene expression / cell surface / magnesium ion binding / ATP binding / identical protein binding / cytosol / cytoplasm
Similarity search - Function
Vinculin-binding site-containing domain / Talin, central / Talin, central domain superfamily / Talin-1/2, rod-segment / : / : / Vinculin Binding Site / Talin, middle domain / Talin, R4 domain / Talin 1-like, rod segment domain ...Vinculin-binding site-containing domain / Talin, central / Talin, central domain superfamily / Talin-1/2, rod-segment / : / : / Vinculin Binding Site / Talin, middle domain / Talin, R4 domain / Talin 1-like, rod segment domain / Talin, N-terminal F0 domain / : / N-terminal or F0 domain of Talin-head FERM / Talin IBS2B domain / I/LWEQ domain / I/LWEQ domain superfamily / I/LWEQ domain / I/LWEQ domain profile. / I/LWEQ domain / Phosphotyrosine-binding domain / IRS-type PTB domain / PTB domain (IRS-1 type) / Alpha-catenin/vinculin-like superfamily / FERM domain signature 1. / FERM conserved site / FERM domain signature 2. / FERM central domain / FERM/acyl-CoA-binding protein superfamily / FERM central domain / FERM superfamily, second domain / FERM domain / FERM domain profile. / Band 4.1 domain / Band 4.1 homologues / 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 / PH-like domain superfamily / Ubiquitin-like domain superfamily
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / Talin-1 / Actin, alpha skeletal muscle
Similarity search - Component
Biological speciesMus musculus (house mouse)
Oryctolagus cuniculus (rabbit)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 2.98 Å
AuthorsBiertumpfel, C. / Yamada, Y. / Mizuno, N.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI) United States
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS) United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2025
Title: Biochemical and structural bases for talin ABSs-F-actin interactions.
Authors: Christian Biertümpfel / Yurika Yamada / Victor Vasquez-Montes / Thien Van Truong / A King Cada / Naoko Mizuno /
Abstract: Focal adhesions (FAs) are large intracellular macromolecular assemblies that play a critical role in cell polarization and migration. Talin serves as a direct connection between integrin receptor and ...Focal adhesions (FAs) are large intracellular macromolecular assemblies that play a critical role in cell polarization and migration. Talin serves as a direct connection between integrin receptor and actomyosin cytoskeleton within FAs. Talin contains three actin-binding sites (ABS1-3) that engage discreetly during the development of FAs, thus acting as a critical player in FA initiation and maturation. However, the molecular basis of the ABS-F-actin interactions remains unknown. Here, we explore interactions of ABSs with F-actin to understand the multivalent behavior of talin. Particularly, the cryo-EM structure of the F-actin-ABS3 complex at 2.9 Å shows ABS3 spanning through two actin monomers along the filament axis, each occupied by the R13 rod subdomain and the DD domain. The dimerization of ABS3 occurs through the DD domain where both protomers interact on the actin surface, and the dimerization of talin to the actin surface is necessary for the engagement to F-actin. The R13 helical bundle is distorted upon binding to F-actin and releases the H1 helix from the rest of the bundle. This phenomenon has also been observed with other tension-sensing proteins like vinculin and α-catenin, highlighting that unfolding is relevant for its force sensing activity. On the contrary, ABS2 (R4R8 subdomains), which is thought to be critical for the maintenance of mature FAs, had multiple F-actin-binding regions within ABS2 and the binding likely occurred by these subdomains running through the surface of F-actin, thus strengthening the interactions upon the maturation of FAs.
History
DepositionMar 10, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Feb 12, 2025Provider: repository / Type: Initial release

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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: Talin-1
G: Talin-1
H: Talin-1
I: Talin-1
J: Talin-1
K: Talin-1
L: Talin-1
M: Talin-1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)420,51323
Polymers418,25513
Non-polymers2,25810
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
Actin, alpha skeletal muscle / Alpha-actin-1


Mass: 41875.633 Da / Num. of mol.: 5 / Source method: isolated from a natural source / Details: post-translationally modified / Source: (natural) Oryctolagus cuniculus (rabbit) / Plasmid details: Hypermol #8101-01 / References: UniProt: P68135
#2: Protein
Talin-1


Mass: 26109.645 Da / Num. of mol.: 8 / Fragment: C-terminal fragment (Domains R13-DD)
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Tln1, Tln / Plasmid: pEC-a-GST-3c / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Gold / References: UniProt: P26039
#3: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: ADP, energy-carrying molecule*YM
#4: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY
Has protein modificationY

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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

Component
IDNameTypeDetailsEntity IDParent-IDSource
1F-actin-Talin(R13-DD) complexCOMPLEXTalin(R13-DD) mixed with polymerized F-actin#1-#20MULTIPLE SOURCES
2Talin(R13-DD)COMPLEX#21RECOMBINANT
3F-actinCOMPLEX#11NATURAL
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Mus musculus (house mouse)10090
33Oryctolagus cuniculus (rabbit)9986
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria) / Plasmid: pEC-a-GST-3c
Buffer solutionpH: 7.5 / Details: pH 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
275 mMKClKCl1
31 mMMgCl2MgCl21
41 mMEGTA[-CH2OCH2CH2N(CH2CO2H)2]21
51 mMDTTHSCH2CH(OH)CH(OH)CH2SH1
SpecimenConc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: G-actin was polymerized for 60 min at RT. 90 uM talin fragments were then incubated with 10 uM F-actin for 30 min at RT and ultra-centrifuged at 100,000 x g for 20 minutes at RT
Specimen supportGrid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 90 % / Chamber temperature: 277 K
Details: 3.0 ul sample, blotted for 5 s from both sides with filter paper Whatman No.1

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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 magnification: 105000 X / Nominal defocus max: 2600 nm / Nominal defocus min: 800 nm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 6.6 sec. / Electron dose: 52 e/Å2 / Film or detector model: GATAN K3 BIOCONTINUUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 6767
EM imaging opticsEnergyfilter name: GIF Bioquantum

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Processing

EM software
IDNameVersionCategory
1RELION3.1.0particle selection
2SerialEMimage acquisition
4CTFFIND4CTF correction
7Coot0.9.8.6model fitting
9RELION3.1.0initial Euler assignment
10RELION3.1.0final Euler assignment
11RELION3.1.0classification
12RELION3.1.03D reconstruction
13PHENIX1.21-5207model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Helical symmerty
IDImage processing-IDAngular rotation/subunit (°)Axial rise/subunit (Å)Axial symmetry
11-166.6627.6C1
21-166.6627.6C1
Particle selectionNum. of particles selected: 2280210
3D reconstructionResolution: 2.98 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 206326 / Algorithm: BACK PROJECTION / Num. of class averages: 1 / Symmetry type: HELICAL
Atomic model buildingProtocol: BACKBONE TRACE / Space: REAL
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
18a2t

8a2t

18a2t1PDBexperimental model
22jsw

2jsw

12jsw2PDBexperimental model
32qdq

2qdq

12qdq3PDBexperimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00422187
ELECTRON MICROSCOPYf_angle_d0.65629981
ELECTRON MICROSCOPYf_dihedral_angle_d6.3883088
ELECTRON MICROSCOPYf_chiral_restr0.0423367
ELECTRON MICROSCOPYf_plane_restr0.0043880

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