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Yorodumi- EMDB-8162: Structure of the F-actin-tropomyosin complex (reprocessed, tropom... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-8162 | |||||||||
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Title | Structure of the F-actin-tropomyosin complex (reprocessed, tropomyosin filtered to 7 Angstrom) | |||||||||
Map data | None | |||||||||
Sample |
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Function / homology | Function and homology information cytoskeletal motor activator activity / tropomyosin binding / mesenchyme migration / troponin I binding / myosin heavy chain binding / filamentous actin / actin filament bundle / skeletal muscle thin filament assembly / striated muscle thin filament / actin filament bundle assembly ...cytoskeletal motor activator activity / tropomyosin binding / mesenchyme migration / troponin I binding / myosin heavy chain binding / filamentous actin / actin filament bundle / skeletal muscle thin filament assembly / striated muscle thin filament / actin filament bundle assembly / 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 | |||||||||
Biological species | Oryctolagus cuniculus (rabbit) / Mus musculus (house mouse) / Rabbit (rabbit) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.6 Å | |||||||||
Authors | von der Ecken J / Raunser S | |||||||||
Funding support | Germany, 2 items
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Citation | Journal: Nature / Year: 2015 Title: Structure of the F-actin-tropomyosin complex. Authors: Julian von der Ecken / Mirco Müller / William Lehman / Dietmar J Manstein / Pawel A Penczek / Stefan Raunser / Abstract: Filamentous actin (F-actin) is the major protein of muscle thin filaments, and actin microfilaments are the main component of the eukaryotic cytoskeleton. Mutations in different actin isoforms lead ...Filamentous actin (F-actin) is the major protein of muscle thin filaments, and actin microfilaments are the main component of the eukaryotic cytoskeleton. Mutations in different actin isoforms lead to early-onset autosomal dominant non-syndromic hearing loss, familial thoracic aortic aneurysms and dissections, and multiple variations of myopathies. In striated muscle fibres, the binding of myosin motors to actin filaments is mainly regulated by tropomyosin and troponin. Tropomyosin also binds to F-actin in smooth muscle and in non-muscle cells and stabilizes and regulates the filaments there in the absence of troponin. Although crystal structures for monomeric actin (G-actin) are available, a high-resolution structure of F-actin is still missing, hampering our understanding of how disease-causing mutations affect the function of thin muscle filaments and microfilaments. Here we report the three-dimensional structure of F-actin at a resolution of 3.7 Å in complex with tropomyosin at a resolution of 6.5 Å, determined by electron cryomicroscopy. The structure reveals that the D-loop is ordered and acts as a central region for hydrophobic and electrostatic interactions that stabilize the F-actin filament. We clearly identify map density corresponding to ADP and Mg(2+) and explain the possible effect of prominent disease-causing mutants. A comparison of F-actin with G-actin reveals the conformational changes during filament formation and identifies the D-loop as their key mediator. We also confirm that negatively charged tropomyosin interacts with a positively charged groove on F-actin. Comparison of the position of tropomyosin in F-actin-tropomyosin with its position in our previously determined F-actin-tropomyosin-myosin structure reveals a myosin-induced transition of tropomyosin. Our results allow us to understand the role of individual mutations in the genesis of actin- and tropomyosin-related diseases and will serve as a strong foundation for the targeted development of drugs. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_8162.map.gz | 27.5 MB | EMDB map data format | |
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Header (meta data) | emd-8162-v30.xml emd-8162.xml | 20.1 KB 20.1 KB | Display Display | EMDB header |
Images | emd_8162.png | 129.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-8162 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-8162 | HTTPS FTP |
-Validation report
Summary document | emd_8162_validation.pdf.gz | 215.3 KB | Display | EMDB validaton report |
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Full document | emd_8162_full_validation.pdf.gz | 214.4 KB | Display | |
Data in XML | emd_8162_validation.xml.gz | 6.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8162 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8162 | HTTPS FTP |
-Related structure data
Related structure data | 5jlfMC 8163C 8164C 8165C 5jlhC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_8162.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | None | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.1 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : F-actin-tropomyosin complex
Entire | Name: F-actin-tropomyosin complex |
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Components |
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-Supramolecule #1: F-actin-tropomyosin complex
Supramolecule | Name: F-actin-tropomyosin complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2 / Details: Filament |
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Molecular weight | Theoretical: 200 KDa |
-Supramolecule #2: F-actin
Supramolecule | Name: F-actin / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1 |
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Source (natural) | Organism: Oryctolagus cuniculus (rabbit) |
-Supramolecule #3: tropomyosin
Supramolecule | Name: tropomyosin / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2 |
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Source (natural) | Organism: Mus musculus (house mouse) |
Recombinant expression | Organism: Escherichia coli (E. coli) |
-Macromolecule #1: Actin, alpha skeletal muscle
Macromolecule | Name: Actin, alpha skeletal muscle / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO |
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Source (natural) | Organism: Rabbit (rabbit) |
Molecular weight | Theoretical: 41.875633 KDa |
Sequence | String: DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IE(HIC)GII TNW DDMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSG DG VTHNVPIYEG ...String: DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IE(HIC)GII TNW DDMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSG DG VTHNVPIYEG YALPHAIMRL DLAGRDLTDY LMKILTERGY SFVTTAEREI VRDIKEKLCY VALDFENEMA TAASSSSL E KSYELPDGQV ITIGNERFRC PETLFQPSFI GMESAGIHET TYNSIMKCDI DIRKDLYANN VMSGGTTMYP GIADRMQKE ITALAPSTMK IKIIAPPERK YSVWIGGSIL ASLSTFQQMW ITKQEYDEAG PSIVHRKCF |
-Macromolecule #2: Tropomyosin Alpha-1
Macromolecule | Name: Tropomyosin Alpha-1 / type: protein_or_peptide / ID: 2 Details: MOUSE TROPOMYSIN WAS USED (UNP P58771, RESIDUES 97-231). DUE TO THE LIMITED RESOLUTION OF THE CRYO-EM DENSITY IN THE REGION OF TROPOMYOSIN, TROPOMYOSIN HAS BEEN REPRESENTED AS POLY(UNK). Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Mus musculus (house mouse) |
Molecular weight | Theoretical: 11.507176 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK) ...String: (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) |
-Macromolecule #3: ADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 5 / Formula: ADP |
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Molecular weight | Theoretical: 427.201 Da |
Chemical component information | ChemComp-ADP: |
-Macromolecule #4: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 5 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | filament |
-Sample preparation
Buffer | pH: 7.5 Component:
Details: 5 mM Tris-HCl pH 7.5, 1 mM DTT, 100 mM KCl, and 2 mM MgCl2 | ||||||||||
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Grid | Model: C-flat-2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE | ||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 90 % / Instrument: GATAN CRYOPLUNGE 3 Details: Sample was applied to a glow-discharged holey carbon grid, incubated for 10 s and manually blotted for 3 s from the backside with filter paper.. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Details | Cs corrected microscope |
Image recording | Film or detector model: FEI FALCON II (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 2-8 / Average exposure time: 0.475 sec. / Average electron dose: 16.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated defocus max: 2.6 µm / Calibrated defocus min: 0.8 µm / Illumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal magnification: 59000 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Initial model | PDB ID: |
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Refinement | Space: RECIPROCAL / Protocol: BACKBONE TRACE / Overall B value: 98 |
Output model | PDB-5jlf: |
-Atomic model buiding 2
Details | tropomyosin fitting |
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Refinement | Protocol: RIGID BODY FIT |
Output model | PDB-5jlf: |