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- EMDB-1481: cryo electron microscopy structure of Vps4p AMPPNP-complexed tetr... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-1481 | |||||||||
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Title | cryo electron microscopy structure of Vps4p AMPPNP-complexed tetradecamer | |||||||||
![]() | This is a 3D reconstruction of a Vps4p AMPPNP-complexed tetradecamer. The N-terminal MIT-domain is deleted. | |||||||||
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Biological species | ![]() ![]() ![]() | |||||||||
Method | ![]() ![]() | |||||||||
![]() | Hartmann C / Chami M / Zachariae U / de Groot BL / Engel A / Gruetter MG | |||||||||
![]() | ![]() Title: Vacuolar protein sorting: two different functional states of the AAA-ATPase Vps4p. Authors: Claudia Hartmann / Mohamed Chami / Ulrich Zachariae / Bert L de Groot / Andreas Engel / Markus G Grütter / ![]() Abstract: The vacuolar protein sorting (Vps) pathway, in which Vps4 class I AAA-ATPases play a central role, regulates growth factor receptors, immune response, and developmental signaling, and participates in ...The vacuolar protein sorting (Vps) pathway, in which Vps4 class I AAA-ATPases play a central role, regulates growth factor receptors, immune response, and developmental signaling, and participates in tumor suppression, apoptosis, and retrovirus budding. We present the first atomic structure of the nucleotide-free yeast His(6)DeltaNVps4p dimer and its AMPPNP (5'-adenylyl-beta,gamma-imidodiphosphate)-bound tetradecamer, derived from a cryo electron microscopy map. Vps4p dimers form two distinct heptameric rings and accommodate AAA cassettes in a head-to-head--not in a head-to-tail-fashion as in class II AAA-ATPases. Our model suggests a mechanism for disassembling ESCRT (endosomal sorting complex required for transport) complexes by movements of substrate-binding domains located at the periphery of the tetradecamer during ATP hydrolysis in one ring, followed by translocation through the central pore and ATP hydrolysis in the second ring. | |||||||||
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 | ![]() | 5 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 9.6 KB 9.6 KB | Display Display | ![]() |
Images | ![]() | 56.4 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
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Links
EMDB pages | ![]() ![]() |
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Map
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | This is a 3D reconstruction of a Vps4p AMPPNP-complexed tetradecamer. The N-terminal MIT-domain is deleted. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : His-tagged Vps4p AAA-ATPase cassette
Entire | Name: His-tagged Vps4p AAA-ATPase cassette |
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Components |
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-Supramolecule #1000: His-tagged Vps4p AAA-ATPase cassette
Supramolecule | Name: His-tagged Vps4p AAA-ATPase cassette / type: sample / ID: 1000 / Details: fresh prepared / Oligomeric state: tetradecamer / Number unique components: 1 |
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Molecular weight | Experimental: 512 KDa / Theoretical: 512 KDa / Method: cryo electron microscopy |
-Macromolecule #1: Vps4p
Macromolecule | Name: Vps4p / type: protein_or_peptide / ID: 1 / Name.synonym: Vps4p / Number of copies: 14 / Oligomeric state: tetradecamer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() ![]() |
Molecular weight | Experimental: 512 KDa / Theoretical: 512 KDa |
Recombinant expression | Organism: ![]() ![]() ![]() |
-Experimental details
-Structure determination
Method | ![]() |
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Aggregation state | particle |
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Sample preparation
Concentration | 2.5 mg/mL |
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Buffer | pH: 7.5 / Details: 20 mM Tris |
Grid | Details: holey carbon film |
Vitrification | Cryogen name: ETHANE / Chamber temperature: 97 K / Instrument: HOMEMADE PLUNGER Details: Vitrification instrument: home made Guillotine. add AMPPNP 15min before freezing. Use the Quantifoil holey carbone grids. Method: Blot for 3 seconds before plunging |
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Electron microscopy
Microscope | FEI/PHILIPS CM200FEG |
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Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 50000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD![]() |
Sample stage | Specimen holder: Cryholder Gatan 626, / Specimen holder model: GATAN LIQUID NITROGEN |
Temperature | Min: 97 K / Max: 97 K / Average: 97 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism corrected at 50 000 Legacy - Electron beam tilt params: no tilt |
Image recording | Category: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: PRIMESCAN / Digitization - Sampling interval: 10 µm / Number real images: 80 / Average electron dose: 10 e/Å2 / Bits/pixel: 8 |
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Image processing
CTF correction | Details: ctftilt, SPIDER |
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Final reconstruction | Applied symmetry - Point group: C7 (7 fold cyclic![]() |
Details | boxer |