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Open data
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Basic information
Entry | Database: PDB / ID: 7bti | |||||||||
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Title | Phalloidin bound F-actin complex | |||||||||
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![]() | CONTRACTILE PROTEIN/PROTEIN BINDING / F-actin / ADP-F-actin / CONTRACTILE PROTEIN / CONTRACTILE PROTEIN-PROTEIN BINDING complex | |||||||||
Function / homology | ![]() Striated Muscle Contraction / skeletal muscle thin filament assembly / striated muscle thin filament / skeletal muscle fiber development / stress fiber / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / toxin activity / hydrolase activity / ATP binding Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() | |||||||||
Method | ELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 3.6 Å | |||||||||
![]() | Kumari, A. / Ragunath, V.K. / Sirajuddin, M. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural insights into actin filament recognition by commonly used cellular actin markers. Authors: Archana Kumari / Shubham Kesarwani / Manjunath G Javoor / Kutti R Vinothkumar / Minhajuddin Sirajuddin / ![]() Abstract: Cellular studies of filamentous actin (F-actin) processes commonly utilize fluorescent versions of toxins, peptides, and proteins that bind actin. While the choice of these markers has been largely ...Cellular studies of filamentous actin (F-actin) processes commonly utilize fluorescent versions of toxins, peptides, and proteins that bind actin. While the choice of these markers has been largely based on availability and ease, there is a severe dearth of structural data for an informed judgment in employing suitable F-actin markers for a particular requirement. Here, we describe the electron cryomicroscopy structures of phalloidin, lifeAct, and utrophin bound to F-actin, providing a comprehensive high-resolution structural comparison of widely used actin markers and their influence towards F-actin. Our results show that phalloidin binding does not induce specific conformational change and lifeAct specifically recognizes closed D-loop conformation, i.e., ADP-Pi or ADP states of F-actin. The structural models aided designing of minimal utrophin and a shorter lifeAct, which can be utilized as F-actin marker. Together, our study provides a structural perspective, where the binding sites of utrophin and lifeAct overlap with majority of actin-binding proteins and thus offering an invaluable resource for researchers in choosing appropriate actin markers and generating new marker variants. | |||||||||
History |
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Structure visualization
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 328.7 KB | Display | ![]() |
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PDB format | ![]() | 270.8 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 30179MC ![]() 6m5gC ![]() 7bt7C ![]() 7bteC C: citing same article ( M: map data used to model this data |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 42096.953 Da / Num. of mol.: 5 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() #2: Protein/peptide | #3: Chemical | ChemComp-MG / #4: Chemical | ChemComp-ADP / Has ligand of interest | Y | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: FILAMENT / 3D reconstruction method: helical reconstruction |
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Sample preparation
Component | Name: Filamentous actin in ADP state / Type: COMPLEX / Entity ID: #1-#2 / Source: NATURAL |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: ![]() ![]() |
Buffer solution | pH: 7.5 Details: 50mM KCl,1mM MgCl2,0.2mM EGTA, 10mM Imidazole buffer pH 7.5 |
Specimen | Conc.: 0.0002 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: 10 mole excess of lifeact was mixed with F-actin and used for sample preparation. |
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK I / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 293 K / Details: blot for 3.5 seconds |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: OTHER / Calibrated magnification: 75000 X / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 120 K / Temperature (min): 100 K |
Image recording | Average exposure time: 2 sec. / Electron dose: 49.2 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 529 / Details: 30 |
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Processing
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EM software |
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CTF correction | Details: GCTF for CTF correction / Type: NONE | ||||||||||||||||||||||||||||||||||||||||||||||||||
Helical symmerty | Angular rotation/subunit: -167 ° / Axial rise/subunit: 27.89 Å / Axial symmetry: C1 | ||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 349839 | ||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 91245 / Num. of class averages: 4 / Symmetry type: HELICAL | ||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 179 / Protocol: RIGID BODY FIT / Space: REAL | ||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 5ONV Pdb chain-ID: A / Pdb chain residue range: 6-373 | ||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement | Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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