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Yorodumi- EMDB-26063: Single-particle Cryo-EM structure of Arp2/3 complex at branched-a... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-26063 | |||||||||
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Title | Single-particle Cryo-EM structure of Arp2/3 complex at branched-actin junction. | |||||||||
Map data | Arp2/3 mediated branched actin junction sharpened map | |||||||||
Sample |
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Function / homology | Function and homology information EPHB-mediated forward signaling / Regulation of actin dynamics for phagocytic cup formation / RHO GTPases Activate WASPs and WAVEs / Arp2/3 protein complex / Arp2/3 complex-mediated actin nucleation / regulation of actin filament polymerization / Clathrin-mediated endocytosis / Neutrophil degranulation / cytoskeletal motor activator activity / tropomyosin binding ...EPHB-mediated forward signaling / Regulation of actin dynamics for phagocytic cup formation / RHO GTPases Activate WASPs and WAVEs / Arp2/3 protein complex / Arp2/3 complex-mediated actin nucleation / regulation of actin filament polymerization / Clathrin-mediated endocytosis / Neutrophil degranulation / cytoskeletal motor activator activity / tropomyosin binding / mesenchyme migration / troponin I binding / myosin heavy chain binding / actin filament bundle / filamentous actin / skeletal muscle thin filament assembly / striated muscle thin filament / actin filament bundle assembly / skeletal muscle myofibril / positive regulation of double-strand break repair via homologous recombination / actin monomer binding / cilium assembly / skeletal muscle fiber development / positive regulation of lamellipodium assembly / stress fiber / titin binding / actin filament polymerization / filopodium / cell projection / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / structural constituent of cytoskeleton / calcium-dependent protein binding / actin filament binding / cell migration / lamellipodium / site of double-strand break / cell body / hydrolase activity / neuron projection / protein domain specific binding / synapse / calcium ion binding / positive regulation of gene expression / magnesium ion binding / positive regulation of transcription by RNA polymerase II / ATP binding / identical protein binding / nucleus / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Bos taurus (cattle) / cattle (cattle) / rabbit (rabbit) / Amanita phalloides (death cap) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.9 Å | |||||||||
Authors | Ding B / Narvaez-Ortiz HY / Nolen BJ / Chowdhury S | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2022 Title: Structure of Arp2/3 complex at a branched actin filament junction resolved by single-particle cryo-electron microscopy. Authors: Bojian Ding / Heidy Y Narvaez-Ortiz / Yuvraj Singh / Glen M Hocky / Saikat Chowdhury / Brad J Nolen / Abstract: Arp2/3 complex nucleates branched actin filaments that provide pushing forces to drive cellular processes such as lamellipodial protrusion and endocytosis. Arp2/3 complex is intrinsically inactive, ...Arp2/3 complex nucleates branched actin filaments that provide pushing forces to drive cellular processes such as lamellipodial protrusion and endocytosis. Arp2/3 complex is intrinsically inactive, and multiple classes of nucleation promoting factors (NPFs) stimulate its nucleation activity. When activated by WASP family NPFs, the complex must bind to the side of a preexisting (mother) filament of actin to complete the nucleation process, ensuring that WASP-mediated activation creates branched rather than linear actin filaments. How actin filaments contribute to activation is currently not understood, largely due to the lack of high-resolution structures of activated Arp2/3 complex bound to the side of a filament. Here, we present the 3.9-Å cryo-electron microscopy structure of the Arp2/3 complex at a branch junction. The structure reveals contacts between Arp2/3 complex and the side of the mother actin filament that likely stimulate subunit flattening, a conformational change that allows the actin-related protein subunits in the complex (Arp2 and Arp3) to mimic filamentous actin subunits. In contrast, limited contact between the bottom half of the complex and the mother filament suggests that clamp twisting, a second major conformational change observed in the active state, is not stimulated by actin filaments, potentially explaining why actin filaments are required but insufficient to trigger nucleation during WASP-mediated activation. Along with biochemical and live-cell imaging data and molecular dynamics simulations, the structure reveals features critical for the interaction of Arp2/3 complex with actin filaments and regulated assembly of branched actin filament networks in cells. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_26063.map.gz | 21.9 MB | EMDB map data format | |
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Header (meta data) | emd-26063-v30.xml emd-26063.xml | 32.6 KB 32.6 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_26063_fsc.xml | 16.8 KB | Display | FSC data file |
Images | emd_26063.png | 100.8 KB | ||
Masks | emd_26063_msk_1.map | 190.1 MB | Mask map | |
Others | emd_26063_additional_1.map.gz emd_26063_additional_2.map.gz emd_26063_half_map_1.map.gz emd_26063_half_map_2.map.gz | 97.2 MB 22.4 MB 179.3 MB 179.3 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-26063 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-26063 | HTTPS FTP |
-Validation report
Summary document | emd_26063_validation.pdf.gz | 498.2 KB | Display | EMDB validaton report |
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Full document | emd_26063_full_validation.pdf.gz | 497.7 KB | Display | |
Data in XML | emd_26063_validation.xml.gz | 20.3 KB | Display | |
Data in CIF | emd_26063_validation.cif.gz | 27.2 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-26063 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-26063 | HTTPS FTP |
-Related structure data
Related structure data | 7tptMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_26063.map.gz / Format: CCP4 / Size: 190.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Arp2/3 mediated branched actin junction sharpened map | ||||||||||||||||||||
Voxel size | X=Y=Z: 1.12 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_26063_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Arp2/3 mediated branched actin junction masked unsharpened map
File | emd_26063_additional_1.map | ||||||||||||
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Annotation | Arp2/3 mediated branched actin junction masked unsharpened map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: Arp2/3 mediated branched actin junction sharpened map with...
File | emd_26063_additional_2.map | ||||||||||||
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Annotation | Arp2/3 mediated branched actin junction sharpened map with peripheral actins | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Arp2/3 mediated branched actin junction masked first half map
File | emd_26063_half_map_1.map | ||||||||||||
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Annotation | Arp2/3 mediated branched actin junction masked first half map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Arp2/3 mediated branched actin junction masked second half map
File | emd_26063_half_map_2.map | ||||||||||||
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Annotation | Arp2/3 mediated branched actin junction masked second half map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : Complex consisting of Arp2/3 complex-mediated branched actin junction
+Supramolecule #1: Complex consisting of Arp2/3 complex-mediated branched actin junction
+Macromolecule #1: Actin-related protein 3
+Macromolecule #2: Actin-related protein 2
+Macromolecule #3: Actin-related protein 2/3 complex subunit 1B
+Macromolecule #4: Actin-related protein 2/3 complex subunit 2
+Macromolecule #5: Actin-related protein 2/3 complex subunit 3
+Macromolecule #6: Actin-related protein 2/3 complex subunit 4
+Macromolecule #7: Actin-related protein 2/3 complex subunit 5
+Macromolecule #8: Actin, alpha skeletal muscle
+Macromolecule #9: Phalloidin
+Macromolecule #10: MAGNESIUM ION
+Macromolecule #11: ADENOSINE-5'-DIPHOSPHATE
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Grid | Model: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.019 kPa / Details: 30 mA |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277.15 K / Instrument: HOMEMADE PLUNGER |
-Electron microscopy
Microscope | FEI TALOS ARCTICA |
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Details | Data were collected by shifting of stage to targeted exposure position. Stage was tilted to different angles: including 40 degree, 36 degree, 30 degree, 25 degree, 15 degree, 0 degree, -20 degree, and -33 degree during data collection. |
Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Digitization - Sampling interval: 14.0 µm / Number grids imaged: 3 / Number real images: 7436 / Average exposure time: 70.0 sec. / Average electron dose: 41.97 e/Å2 Details: Each micrograph was acquired as dose-fractionated movies consisting of 82 frames per movie. |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.2 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 92000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: FLEXIBLE FIT |
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Output model | PDB-7tpt: |