|Entry||Database: PDB / ID: 3j95|
|Title||Structure of ADP-bound N-ethylmaleimide sensitive factor determined by single particle cryoelectron microscopy|
|Keywords||HYDROLASE / ATPases associated with diverse cellular activities|
|Function / homology|
Function and homology information
SNARE complex disassembly / vesicle-fusing ATPase / positive regulation of receptor recycling / syntaxin-1 binding / ionotropic glutamate receptor binding / SNARE binding / potassium ion transport / ATPase activity, coupled / PDZ domain binding / positive regulation of protein catabolic process ...SNARE complex disassembly / vesicle-fusing ATPase / positive regulation of receptor recycling / syntaxin-1 binding / ionotropic glutamate receptor binding / SNARE binding / potassium ion transport / ATPase activity, coupled / PDZ domain binding / positive regulation of protein catabolic process / intracellular protein transport / midbody / ATPase activity / protein-containing complex binding / protein kinase binding / ATP binding / identical protein binding / plasma membrane / metal ion binding / cytosol
AAA+ lid domain / CDC48, N-terminal subdomain / Cell division protein 48 (CDC48), domain 2 / Cell division protein 48 (CDC48), N-terminal domain / ATPase family associated with various cellular activities (AAA) / AAA ATPase, AAA+ lid domain / Vesicle-fusing ATPase / CDC48 domain 2-like superfamily / P-loop containing nucleoside triphosphate hydrolase / Aspartate decarboxylase-like domain superfamily ...AAA+ lid domain / CDC48, N-terminal subdomain / Cell division protein 48 (CDC48), domain 2 / Cell division protein 48 (CDC48), N-terminal domain / ATPase family associated with various cellular activities (AAA) / AAA ATPase, AAA+ lid domain / Vesicle-fusing ATPase / CDC48 domain 2-like superfamily / P-loop containing nucleoside triphosphate hydrolase / Aspartate decarboxylase-like domain superfamily / CDC48, domain 2 / ATPase, AAA-type, conserved site / ATPase, AAA-type, core / AAA+ ATPase domain
|Biological species||Cricetulus griseus (Chinese hamster)|
|Method||ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 7.6 Å|
|Authors||Zhao, M. / Wu, S. / Cheng, Y. / Brunger, A.T.|
|Citation||Journal: Nature / Year: 2015|
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.
SummaryFull reportAbout validation report
|Structure viewer||Molecule: |
Downloads & links
A: Vesicle-fusing ATPase
B: Vesicle-fusing ATPase
C: Vesicle-fusing ATPase
D: Vesicle-fusing ATPase
E: Vesicle-fusing ATPase
F: Vesicle-fusing ATPase
Mass: 82907.430 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Cricetulus griseus (Chinese hamster) / Gene: NSF / Production host: Escherichia coli (E. coli) / References: UniProt: P18708, vesicle-fusing ATPase
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction|
|Component||Name: ADP-bound N-ethylmaleimide sensitive factor / Type: COMPLEX / Details: hexamer|
|Molecular weight||Value: 0.5 MDa / Experimental value: NO|
|Buffer solution||Name: 50 mM Tris-Cl, 150 mM NaCl, 1 mM EDTA, 1 mM ATP, 1 mM DTT, 0.05% v/v Nonident P-40|
Details: 50 mM Tris-Cl, 150 mM NaCl, 1 mM EDTA, 1 mM ATP, 1 mM DTT, 0.05% v/v Nonident P-40
|Specimen||Conc.: 15 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Details: Holey carbon on top of 400 mesh copper grid|
|Vitrification||Instrument: FEI VITROBOT MARK I / Cryogen name: ETHANE / Temp: 90 K / Humidity: 100 %|
Details: Blot for 3.5 seconds before plunging into liquid ethane (FEI VITROBOT MARK I).
Method: Blot for 3.5 seconds before plunging
-Electron microscopy imaging
Model: Tecnai Polara / Image courtesy: FEI Company
|Microscopy||Model: FEI POLARA 300 / Date: Jan 14, 2014|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 31000 X / Nominal defocus max: -2800 nm / Nominal defocus min: -1800 nm / Cs: 2.3 mm / Camera length: 0 mm|
|Specimen holder||Model: OTHER / Specimen holder type: unspecified|
|Image recording||Electron dose: 44 e/Å2 / Film or detector model: GATAN K2 (4k x 4k)|
|Radiation||Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray|
|Radiation wavelength||Relative weight: 1|
|CTF correction||Details: Each particle|
|Symmetry||Point symmetry: C1 (asymmetric)|
|3D reconstruction||Resolution: 7.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 12830 / Nominal pixel size: 2.4312 Å / Actual pixel size: 2.4312 Å / Details: (Single particle--Applied symmetry: C1) / Symmetry type: POINT|
|Atomic model building||Protocol: FLEXIBLE FIT / Space: RECIPROCAL / Target criteria: R-factor|
Details: REFINEMENT PROTOCOL--flexible DETAILS--D2 domain of NSF was from crystal structure 1NSF. D1 domain of NSF was from related entry EMD-6204.
|Atomic model building||PDB-ID: 1NSF|
Pdb chain-ID: A
|Refinement||Resolution: 7.601→311.194 Å / SU ML: 1.3 / σ(F): 0.29 / Phase error: 36.78 / Stereochemistry target values: ML|
|Solvent computation||Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL|
|Displacement parameters||Biso max: 50 Å2 / Biso mean: 50 Å2 / Biso min: 50 Å2|
|Refinement step||Cycle: LAST / Resolution: 7.601→311.194 Å|
|Refine LS restraints|
|LS refinement shell|
Refinement-ID: ELECTRON MICROSCOPY / Total num. of bins used: 30
-Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)
EMDB accession codes are about to change! (news from PDBe EMDB page)
- The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force. (see PDBe EMDB page)
- The EM Navigator/Yorodumi systems omit the EMD- prefix.
Related info.:Q: What is "EMD"? / ID/Accession-code notation in Yorodumi/EM Navigator
-Jul 12, 2017. Major update of PDB
Major update of PDB
- wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary. This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated. See below links for details.
- In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software). Now, EM Navigator and Yorodumi are based on the updated data.
+Jun 16, 2017. Omokage search with filter
Omokage search with filter
- Result of Omokage search can be filtered by keywords and the database types
Related info.:Omokage search
+Sep 15, 2016. EM Navigator & Yorodumi renewed
EM Navigator & Yorodumi renewed
- New versions of EM Navigator and Yorodumi started
Related info.:Changes in new EM Navigator and Yorodumi
+Aug 31, 2016. New EM Navigator & Yorodumi
New EM Navigator & Yorodumi
- In 15th Sep 2016, the development versions of EM Navigator and Yorodumi will replace the official versions.
- Current version will continue as 'legacy version' for some time.
Related info.:Changes in new EM Navigator and Yorodumi / EM Navigator / Yorodumi
Thousand views of thousand structures
- Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
- This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi