|Entry||Database: PDB / ID: 6mdo|
|Title||The D1 and D2 domain rings of NSF engaging the SNAP-25 N-terminus within the 20S supercomplex (focused refinement on D1/D2 rings, class 1)|
|Keywords||HYDROLASE / SNARE / NSF / SNAP / ATPase / AAA / disassembly / synapse / membrane fusion / exocytosis|
|Function / homology||AAA+ lid domain / P-loop containing nucleoside triphosphate hydrolase / Target SNARE coiled-coil homology domain / SNAP-25 / CDC48, N-terminal subdomain / ATPase, AAA-type, core / ATPase, AAA-type, conserved site / CDC48, domain 2 / Aspartate decarboxylase-like domain superfamily / AAA+ ATPase domain ...AAA+ lid domain / P-loop containing nucleoside triphosphate hydrolase / Target SNARE coiled-coil homology domain / SNAP-25 / CDC48, N-terminal subdomain / ATPase, AAA-type, core / ATPase, AAA-type, conserved site / CDC48, domain 2 / Aspartate decarboxylase-like domain superfamily / AAA+ ATPase domain / CDC48 domain 2-like superfamily / Vesicle-fusing ATPase / ATPase family associated with various cellular activities (AAA) / SNAP-25 family / Cell division protein 48 (CDC48), N-terminal domain / Cell division protein 48 (CDC48), domain 2 / AAA-protein family signature. / t-SNARE coiled-coil homology domain profile. / Synaptosomal-associated protein 25 / SNARE complex disassembly / anchored component of presynaptic membrane / growth hormone secretion / synaptobrevin 2-SNAP-25-syntaxin-1a complex / synaptobrevin 2-SNAP-25-syntaxin-1a-complexin II complex / synaptobrevin 2-SNAP-25-syntaxin-1a-complexin I complex / presynaptic active zone membrane / synaptic vesicle fusion to presynaptic active zone membrane / SNARE complex assembly / synaptic vesicle priming / intracellular organelle / calcium ion-regulated exocytosis of neurotransmitter / regulation of synapse assembly / vesicle-fusing ATPase / vesicle fusion / positive regulation of hormone secretion / SNAP receptor activity / sleep / synaptic vesicle exocytosis / regulation of neuron projection development / SNARE complex / positive regulation of receptor recycling / syntaxin-1 binding / myosin binding / syntaxin binding / voltage-gated potassium channel activity / exocytosis / ionotropic glutamate receptor binding / long-term memory / positive regulation of insulin secretion / SNARE binding / voltage-gated potassium channel complex / axonal growth cone / axonogenesis / filopodium / endosomal transport / potassium ion transport / ATPase activity, coupled / neuron differentiation / PDZ domain binding / positive regulation of protein catabolic process / terminal bouton / calcium-dependent protein binding / intracellular protein transport / actin cytoskeleton / midbody / growth cone / lamellipodium / protein N-terminus binding / ion channel binding / ATPase activity / endosome / protein-containing complex binding / cell junction / neuron projection / membrane raft / myelin sheath / axon / glutamatergic synapse / protein domain specific binding / neuronal cell body / protein kinase binding / perinuclear region of cytoplasm / membrane / ATP binding / identical protein binding / plasma membrane / metal ion binding / cytosol / cytoplasm / Vesicle-fusing ATPase / Synaptosomal-associated protein 25|
Function and homology information
|Specimen source||Cricetulus griseus (Chinese hamster)|
Rattus norvegicus (Norway rat)
|Method||ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / 3.9 Å resolution|
|Authors||White, K.I. / Zhao, M. / Brunger, A.T.|
|Citation||Journal: Elife / Year: 2018|
Title: Structural principles of SNARE complex recognition by the AAA+ protein NSF.
Authors: K Ian White / Minglei Zhao / Ucheor B Choi / Richard A Pfuetzner / Axel T Brunger
Abstract: The recycling of SNARE proteins following complex formation and membrane fusion is an essential process in eukaryotic trafficking. A highly conserved AAA+ protein, NSF (-ethylmaleimide sensitive ...The recycling of SNARE proteins following complex formation and membrane fusion is an essential process in eukaryotic trafficking. A highly conserved AAA+ protein, NSF (-ethylmaleimide sensitive factor) and an adaptor protein, SNAP (soluble NSF attachment protein), disassemble the SNARE complex. We report electron-cryomicroscopy structures of the complex of NSF, αSNAP, and the full-length soluble neuronal SNARE complex (composed of syntaxin-1A, synaptobrevin-2, SNAP-25A) in the presence of ATP under non-hydrolyzing conditions at ~3.9 Å resolution. These structures reveal electrostatic interactions by which two αSNAP molecules interface with a specific surface of the SNARE complex. This interaction positions the SNAREs such that the 15 N-terminal residues of SNAP-25A are loaded into the D1 ring pore of NSF via a spiral pattern of interactions between a conserved tyrosine NSF residue and SNAP-25A backbone atoms. This loading process likely precedes ATP hydrolysis. Subsequent ATP hydrolysis then drives complete disassembly.
SummaryFull reportAbout validation report
|Date||Deposition: Sep 4, 2018 / Release: Sep 19, 2018|
|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
H: Synaptosomal-associated protein 25
Mass: 85509.227 Da / Num. of mol.: 6 / Source: (gene. exp.) Cricetulus griseus (Chinese hamster) / Gene: NSF / Production host: Escherichia coli (E. coli) / References: UniProt: P18708, vesicle-fusing ATPase
Mass: 23512.387 Da / Num. of mol.: 1 / Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Snap25, Snap / Production host: Escherichia coli (E. coli) / References: UniProt: P60881
Mass: 507.181 Da / Num. of mol.: 9 / Formula: C10H16N5O13P3 / Adenosine triphosphate / Comment: ATP (energy-carrying molecule) *YM
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / Reconstruction method: single particle reconstruction|
|Buffer solution||pH: 8|
|Specimen||Conc.: 15 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Grid material: COPPER / Grid mesh size: 200 / Grid type: Quantifoil R1.2/1.3|
|Vitrification||Instrument: FEI VITROBOT MARK I / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 293 kelvins / Details: Blot for 3.5 seconds before plunging.|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Microscopy||Microscope model: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3000 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm / C2 aperture diameter: 70 microns / Alignment procedure: COMA FREE|
|Specimen holder||Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER|
|Image recording||Average exposure time: 10 sec. / Electron dose: 58 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number of grids imaged: 2 / Number of real images: 5418|
|Image scans||Movie frames/image: 40 / Used frames/image: 2-40|
|CTF correction||Details: CTF correction was carried out in Relion with reconstruction step.|
Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
|Particle selection||Number of particles selected: 475680|
|Symmetry||Point symmetry: C1|
|3D reconstruction||Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 166620 / Algorithm: FOURIER SPACE / Number of class averages: 1 / Symmetry type: POINT|
|Atomic model building||Ref protocol: RIGID BODY FIT / Ref space: REAL|
|Atomic model building||PDB-ID: 3J96|
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