+Open data
-Basic information
Entry | Database: PDB / ID: 6w18 | ||||||
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Title | Structure of S. pombe Arp2/3 complex in inactive state | ||||||
Components | (Actin-related protein ...) x 7 | ||||||
Keywords | STRUCTURAL PROTEIN / Arp2/3 / actin / cytoskeletal protein / actin regulator | ||||||
Function / homology | Function and homology information Regulation of actin dynamics for phagocytic cup formation / RHO GTPases Activate WASPs and WAVEs / Clathrin-mediated endocytosis / actin cortical patch organization / Neutrophil degranulation / medial cortex / cell cortex of cell tip / actin cortical patch assembly / Arp2/3 protein complex / Arp2/3 complex-mediated actin nucleation ...Regulation of actin dynamics for phagocytic cup formation / RHO GTPases Activate WASPs and WAVEs / Clathrin-mediated endocytosis / actin cortical patch organization / Neutrophil degranulation / medial cortex / cell cortex of cell tip / actin cortical patch assembly / Arp2/3 protein complex / Arp2/3 complex-mediated actin nucleation / cell tip / actin cortical patch / regulation of actin filament polymerization / mating projection tip / cortical actin cytoskeleton organization / establishment or maintenance of cell polarity / cell division site / mitotic cytokinesis / actin filament polymerization / structural constituent of cytoskeleton / actin filament binding / endocytosis / cell cortex / ATP binding / nucleus / cytoplasm / cytosol Similarity search - Function | ||||||
Biological species | Schizosaccharomyces pombe (fission yeast) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.2 Å | ||||||
Authors | Shaaban, M. / Nolen, B.J. / Chowdhury, S. | ||||||
Funding support | United States, 1items
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Citation | Journal: Nat Struct Mol Biol / Year: 2020 Title: Cryo-EM reveals the transition of Arp2/3 complex from inactive to nucleation-competent state. Authors: Mohammed Shaaban / Saikat Chowdhury / Brad J Nolen / Abstract: Arp2/3 complex, a crucial actin filament nucleator, undergoes structural rearrangements during activation by nucleation-promoting factors (NPFs). However, the conformational pathway leading to the ...Arp2/3 complex, a crucial actin filament nucleator, undergoes structural rearrangements during activation by nucleation-promoting factors (NPFs). However, the conformational pathway leading to the nucleation-competent state is unclear due to lack of high-resolution structures of the activated state. Here we report a ~3.9 Å resolution cryo-EM structure of activated Schizosaccharomyces pombe Arp2/3 complex bound to the S. pombe NPF Dip1 and attached to the end of the nucleated actin filament. The structure reveals global and local conformational changes that allow the two actin-related proteins in Arp2/3 complex to mimic a filamentous actin dimer and template nucleation. Activation occurs through a clamp-twisting mechanism, in which Dip1 forces two core subunits in Arp2/3 complex to pivot around one another, shifting half of the complex into a new activated position. By showing how Dip1 stimulates activation, the structure reveals how NPFs can activate Arp2/3 complex in diverse cellular processes. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6w18.cif.gz | 327.9 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6w18.ent.gz | 245.9 KB | Display | PDB format |
PDBx/mmJSON format | 6w18.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6w18_validation.pdf.gz | 1.5 MB | Display | wwPDB validaton report |
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Full document | 6w18_full_validation.pdf.gz | 1.5 MB | Display | |
Data in XML | 6w18_validation.xml.gz | 57.9 KB | Display | |
Data in CIF | 6w18_validation.cif.gz | 89.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/w1/6w18 ftp://data.pdbj.org/pub/pdb/validation_reports/w1/6w18 | HTTPS FTP |
-Related structure data
Related structure data | 21503MC 6w17C C: citing same article (ref.) M: map data used to model this data |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Actin-related protein ... , 7 types, 7 molecules ABCDEFG
#1: Protein | Mass: 47427.137 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Schizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast) Plasmid details: Fission yeast / Strain: 972 / ATCC 24843 / References: UniProt: P32390 |
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#2: Protein | Mass: 44286.758 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Schizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast) Plasmid details: Fission yeast / Strain: 972 / ATCC 24843 / References: UniProt: Q9UUJ1 |
#3: Protein | Mass: 41643.465 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Schizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast) Plasmid details: Fission yeast / Strain: 972 / ATCC 24843 / References: UniProt: P78774 |
#4: Protein | Mass: 37025.230 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Schizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast) Plasmid details: Fission yeast / Strain: 972 / ATCC 24843 / References: UniProt: O14241 |
#5: Protein | Mass: 19865.746 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Schizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast) Plasmid details: Fission yeast / Strain: 972 / ATCC 24843 / References: UniProt: Q9Y7J4 |
#6: Protein | Mass: 19637.695 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Schizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast) Strain: 972 / ATCC 24843 / References: UniProt: Q92352 |
#7: Protein | Mass: 16922.059 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Schizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast) Plasmid details: Fission yeast / Strain: 972 / ATCC 24843 / References: UniProt: Q10316 |
-Non-polymers , 2 types, 3 molecules
#8: Chemical | #9: Chemical | ChemComp-MG / | |
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-Details
Has ligand of interest | N |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Arp2/3 complex / Type: COMPLEX / Details: Inactive state of S. pombe Arp2/3 complex / Entity ID: #1-#7 / Source: NATURAL | ||||||||||||||||||||||||||||||
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Molecular weight | Value: 0.225 MDa / Experimental value: NO | ||||||||||||||||||||||||||||||
Source (natural) | Organism: Schizosaccharomyces pombe (strain 972 / ATCC 24843) (yeast) Strain: 972 / ATCC 24843 / Cellular location: cytoplasm | ||||||||||||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 2.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||
Specimen support | Details: 20mA current / Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil, UltrAuFoil, R1.2/1.3 | ||||||||||||||||||||||||||||||
Vitrification | Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 98 % / Chamber temperature: 277.15 K Details: 4 uL of sample was applied to freshly glow-discharged grid. Excess sample was manually blotted off with a dry Whatman No.1 filter paper for 5-7 seconds. Immediately after the blotting step ...Details: 4 uL of sample was applied to freshly glow-discharged grid. Excess sample was manually blotted off with a dry Whatman No.1 filter paper for 5-7 seconds. Immediately after the blotting step the sample containing grid was rapidly vitrified by plunge freezing into liquid ethane. |
-Electron microscopy imaging
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
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Microscopy | Model: FEI TALOS ARCTICA Details: Data were collected by combining untilted and tilted (20, 30 and 40 degrees) images. Stage shifting to the targeted exposure position was used for navigation during data acquisition. |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 120000 X / Nominal defocus max: -1300 nm / Nominal defocus min: -700 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: BASIC |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 40 sec. / Electron dose: 44.34 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 3 / Num. of real images: 5309 Details: Each micrograph was collected as dose-fractionated movies consisting of 62 fractions per movie. |
Image scans | Sampling size: 14 µm / Width: 4000 / Height: 4000 |
-Processing
EM software |
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CTF correction | Details: Local CTF estimation was performed for all micrographs using the CTF estimation package within cryoSPARC. Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 3336981 / Details: Automated Gaussian-based blob picker was used. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 112170 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 3DWL Accession code: 3DWL / Source name: PDB / Type: experimental model | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement | Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 51.2 Å2 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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