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- EMDB-6210: Structure of 20S supercomplex with V7-SNARE determined by single ... -
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Basic information
Entry | Database: EMDB / ID: EMD-6210 | |||||||||
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Title | Structure of 20S supercomplex with V7-SNARE determined by single particle cryoelectron microscopy | |||||||||
![]() | Map of 20S supercomplex with V7-SNARE. This map is unsharpened and unfiltered. The map was normalized using the program MAPMAN. | |||||||||
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![]() | vesicle trafficking | |||||||||
Function / homology | ![]() regulation of protein targeting to vacuolar membrane / positive regulation of histamine secretion by mast cell / : / soluble NSF attachment protein activity / Intra-Golgi traffic / Retrograde transport at the Trans-Golgi-Network / triglyceride transport / natural killer cell degranulation / COPI-dependent Golgi-to-ER retrograde traffic / COPI-mediated anterograde transport ...regulation of protein targeting to vacuolar membrane / positive regulation of histamine secretion by mast cell / : / soluble NSF attachment protein activity / Intra-Golgi traffic / Retrograde transport at the Trans-Golgi-Network / triglyceride transport / natural killer cell degranulation / COPI-dependent Golgi-to-ER retrograde traffic / COPI-mediated anterograde transport / trans-Golgi Network Vesicle Budding / BLOC-1 complex / SNARE complex disassembly / regulation of delayed rectifier potassium channel activity / vesicle fusion with Golgi apparatus / myosin head/neck binding / exocytic insertion of neurotransmitter receptor to postsynaptic membrane / Other interleukin signaling / extrinsic component of presynaptic membrane / synaptobrevin 2-SNAP-25-syntaxin-1a-complexin II complex / synaptobrevin 2-SNAP-25-syntaxin-1a complex / synaptobrevin 2-SNAP-25-syntaxin-1a-complexin I complex / Lysosome Vesicle Biogenesis / synaptic vesicle fusion to presynaptic active zone membrane / regulation of synaptic vesicle priming / Glutamate Neurotransmitter Release Cycle / positive regulation of norepinephrine secretion / neutrophil degranulation / Norepinephrine Neurotransmitter Release Cycle / COPII-mediated vesicle transport / protein-containing complex disassembly / Acetylcholine Neurotransmitter Release Cycle / positive regulation of catecholamine secretion / zymogen granule membrane / Serotonin Neurotransmitter Release Cycle / GABA synthesis, release, reuptake and degradation / Dopamine Neurotransmitter Release Cycle / Golgi Associated Vesicle Biogenesis / regulated exocytosis / presynaptic dense core vesicle exocytosis / ribbon synapse / synaptic vesicle docking / regulation of establishment of protein localization / storage vacuole / response to gravity / calcium ion-regulated exocytosis of neurotransmitter / Insertion of tail-anchored proteins into the endoplasmic reticulum membrane / eosinophil degranulation / vesicle fusion / vesicle docking / positive regulation of calcium ion-dependent exocytosis / chloride channel inhibitor activity / secretion by cell / ATP-dependent protein disaggregase activity / vesicle transport along microtubule / SNARE complex / SNAP receptor activity / Cargo recognition for clathrin-mediated endocytosis / regulation of vesicle-mediated transport / Clathrin-mediated endocytosis / LGI-ADAM interactions / hormone secretion / calcium-ion regulated exocytosis / actomyosin / apical protein localization / positive regulation of intracellular protein transport / intra-Golgi vesicle-mediated transport / Golgi to plasma membrane protein transport / platelet alpha granule / regulation of exocytosis / positive regulation of hormone secretion / neurotransmitter secretion / Golgi stack / neurotransmitter receptor internalization / protein localization to membrane / ATP-dependent protein binding / neuron projection terminus / positive regulation of ATP-dependent activity / vesicle-fusing ATPase / neurotransmitter transport / regulation of synaptic vesicle recycling / positive regulation of dendrite morphogenesis / insulin secretion / syntaxin-1 binding / SNARE complex assembly / positive regulation of neurotransmitter secretion / syntaxin binding / vacuolar membrane / clathrin-coated vesicle / synaptic vesicle priming / Neutrophil degranulation / azurophil granule membrane / regulation of synapse assembly / regulation of neuron projection development / endosomal transport / phagocytosis, engulfment / myosin binding / positive regulation of receptor recycling / endosome to lysosome transport / exocytosis Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 8.0 Å | |||||||||
![]() | Zhao M / Wu S / Zhou Q / Vivona S / Cipriano DJ / Cheng Y / Brunger AT | |||||||||
![]() | ![]() Title: Mechanistic insights into the recycling machine of the SNARE complex. Authors: Minglei Zhao / Shenping Wu / Qiangjun Zhou / Sandro Vivona / Daniel J Cipriano / Yifan Cheng / Axel T Brunger / ![]() Abstract: Evolutionarily conserved SNARE (soluble N-ethylmaleimide sensitive factor attachment protein receptors) proteins form a complex that drives membrane fusion in eukaryotes. The ATPase NSF (N- ...Evolutionarily conserved SNARE (soluble N-ethylmaleimide sensitive factor attachment protein receptors) proteins form a complex that drives membrane fusion in eukaryotes. The ATPase NSF (N-ethylmaleimide sensitive factor), together with SNAPs (soluble NSF attachment protein), disassembles the SNARE complex into its protein components, making individual SNAREs available for subsequent rounds of fusion. Here we report structures of ATP- and ADP-bound NSF, and the NSF/SNAP/SNARE (20S) supercomplex determined by single-particle electron cryomicroscopy at near-atomic to sub-nanometre resolution without imposing symmetry. Large, potentially force-generating, conformational differences exist between ATP- and ADP-bound NSF. The 20S supercomplex exhibits broken symmetry, transitioning from six-fold symmetry of the NSF ATPase domains to pseudo four-fold symmetry of the SNARE complex. SNAPs interact with the SNARE complex with an opposite structural twist, suggesting an unwinding mechanism. The interfaces between NSF, SNAPs, and SNAREs exhibit characteristic electrostatic patterns, suggesting how one NSF/SNAP species can act on many different SNARE complexes. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 6 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 14 KB 14 KB | Display Display | ![]() |
Images | ![]() | 91.4 KB | ||
Others | ![]() | 7.3 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 78.9 KB | Display | ![]() |
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Full document | ![]() | 78 KB | Display | |
Data in XML | ![]() | 493 B | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6204C ![]() 6205C ![]() 6206C ![]() 6207C ![]() 6208C ![]() 6209C ![]() 3j94C ![]() 3j95C ![]() 3j96C ![]() 3j97C ![]() 3j98C ![]() 3j99C C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Map of 20S supercomplex with V7-SNARE. This map is unsharpened and unfiltered. The map was normalized using the program MAPMAN. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.4312 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Supplemental map: emd 6210 additional 1.map
File | emd_6210_additional_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
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Sample components
-Entire : 20S supercomplex consisting of VAMP-7 SNARE complex, alpha-SNAP, ...
Entire | Name: 20S supercomplex consisting of VAMP-7 SNARE complex, alpha-SNAP, and N-ethylmaleimide sensitive factor (NSF) |
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Components |
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-Supramolecule #1000: 20S supercomplex consisting of VAMP-7 SNARE complex, alpha-SNAP, ...
Supramolecule | Name: 20S supercomplex consisting of VAMP-7 SNARE complex, alpha-SNAP, and N-ethylmaleimide sensitive factor (NSF) type: sample / ID: 1000 Oligomeric state: One hexamer of NSF + four alpha-SNAP molecules + one SNARE complex Number unique components: 5 |
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-Macromolecule #1: N-ethylmaleimide sensitive factor
Macromolecule | Name: N-ethylmaleimide sensitive factor / type: protein_or_peptide / ID: 1 / Name.synonym: NSF / Number of copies: 6 / Oligomeric state: hexamer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 83 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | UniProtKB: Vesicle-fusing ATPase |
-Macromolecule #2: alpha Soluble NSF Attachment Protein
Macromolecule | Name: alpha Soluble NSF Attachment Protein / type: protein_or_peptide / ID: 2 / Name.synonym: alpha-SNAP / Number of copies: 4 / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 33 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | UniProtKB: Alpha-soluble NSF attachment protein |
-Macromolecule #3: Syntaxin-1A
Macromolecule | Name: Syntaxin-1A / type: protein_or_peptide / ID: 3 / Name.synonym: Stx-1A / Number of copies: 1 / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 30 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | UniProtKB: Syntaxin-1A |
-Macromolecule #4: Vesicle-associated membrane protein 7
Macromolecule | Name: Vesicle-associated membrane protein 7 / type: protein_or_peptide / ID: 4 / Name.synonym: VAMP-7 / Number of copies: 1 / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 22 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | UniProtKB: Vesicle-associated membrane protein 7 |
-Macromolecule #5: Synaptosomal-associated protein 25
Macromolecule | Name: Synaptosomal-associated protein 25 / type: protein_or_peptide / ID: 5 / Name.synonym: SNAP-25 / Number of copies: 1 / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 25 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | UniProtKB: Synaptosomal-associated protein 25 |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 15 mg/mL |
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Buffer | pH: 8 Details: 50 mM Tris-Cl, 150 mM NaCl, 1 mM ATP, 1 mM EDTA, 1 mM DTT, 0.05% v/v Nonident P-40 |
Grid | Details: Holey carbon on top of 400 mesh copper grid |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 90 K / Instrument: FEI VITROBOT MARK I / Method: Blot for 3.5 seconds before plunging. |
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Electron microscopy
Microscope | FEI POLARA 300 |
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Date | May 12, 2014 |
Image recording | Category: CCD / Film or detector model: GATAN K2 (4k x 4k) / Average electron dose: 44 e/Å2 Details: Gatan K2 Summit in super-resolution counting mode. Motion correction as described in Li et al. (2013) Nature Methods. |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.3 mm / Nominal defocus max: -2.8 µm / Nominal defocus min: -1.8 µm / Nominal magnification: 31000 |
Sample stage | Specimen holder model: OTHER |
Experimental equipment | ![]() Model: Tecnai Polara / Image courtesy: FEI Company |
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Image processing
Details | 3D classification, refinement, and reconstruction were performed using RELION. |
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CTF correction | Details: Each particle |
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 8.0 Å / Resolution method: OTHER / Software - Name: RELION / Number images used: 32100 |