+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-8363 | |||||||||
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Title | CryoEM map of the S. cerevisiae Vo region lacking subunit d | |||||||||
Map data | The Cryo-EM map of yeast Vo lacking the d subunit | |||||||||
Sample |
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Function / homology | Function and homology information cell wall mannoprotein biosynthetic process / ATPase-coupled ion transmembrane transporter activity / cellular response to alkaline pH / Insulin receptor recycling / Transferrin endocytosis and recycling / polyphosphate metabolic process / ROS and RNS production in phagocytes / Amino acids regulate mTORC1 / Golgi lumen acidification / P-type proton-exporting transporter activity ...cell wall mannoprotein biosynthetic process / ATPase-coupled ion transmembrane transporter activity / cellular response to alkaline pH / Insulin receptor recycling / Transferrin endocytosis and recycling / polyphosphate metabolic process / ROS and RNS production in phagocytes / Amino acids regulate mTORC1 / Golgi lumen acidification / P-type proton-exporting transporter activity / plasma membrane proton-transporting V-type ATPase complex / endosomal lumen acidification / vacuolar proton-transporting V-type ATPase, V0 domain / vacuolar transport / vacuole organization / protein targeting to vacuole / proton-transporting V-type ATPase complex / fungal-type vacuole / cellular hyperosmotic response / fungal-type vacuole membrane / vacuolar proton-transporting V-type ATPase complex / phosphatidylinositol-3,5-bisphosphate binding / vacuolar acidification / proton transmembrane transporter activity / intracellular copper ion homeostasis / proton transmembrane transport / Neutrophil degranulation / proton-transporting ATPase activity, rotational mechanism / cell periphery / transmembrane transport / endocytosis / ATPase binding / protein-containing complex assembly / intracellular iron ion homeostasis / early endosome / Golgi membrane / membrane / plasma membrane Similarity search - Function | |||||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 7.8 Å | |||||||||
Authors | Mazhab-Jafari MT / Rohou A / Schmidt C / Bueler SA / Benlekbir S / Robinson C / Rubinstein JL | |||||||||
Citation | Journal: Nature / Year: 2016 Title: Atomic model for the membrane-embedded V motor of a eukaryotic V-ATPase. Authors: Mohammad T Mazhab-Jafari / Alexis Rohou / Carla Schmidt / Stephanie A Bueler / Samir Benlekbir / Carol V Robinson / John L Rubinstein / Abstract: Vacuolar-type ATPases (V-ATPases) are ATP-powered proton pumps involved in processes such as endocytosis, lysosomal degradation, secondary transport, TOR signalling, and osteoclast and kidney ...Vacuolar-type ATPases (V-ATPases) are ATP-powered proton pumps involved in processes such as endocytosis, lysosomal degradation, secondary transport, TOR signalling, and osteoclast and kidney function. ATP hydrolysis in the soluble catalytic V region drives proton translocation through the membrane-embedded V region via rotation of a rotor subcomplex. Variability in the structure of the intact enzyme has prevented construction of an atomic model for the membrane-embedded motor of any rotary ATPase. We induced dissociation and auto-inhibition of the V and V regions of the V-ATPase by starving the yeast Saccharomyces cerevisiae, allowing us to obtain a ~3.9-Å resolution electron cryomicroscopy map of the V complex and build atomic models for the majority of its subunits. The analysis reveals the structures of subunits acc'c″de and a protein that we identify and propose to be a new subunit (subunit f). A large cavity between subunit a and the c-ring creates a cytoplasmic half-channel for protons. The c-ring has an asymmetric distribution of proton-carrying Glu residues, with the Glu residue of subunit c″ interacting with Arg735 of subunit a. The structure suggests sequential protonation and deprotonation of the c-ring, with ATP-hydrolysis-driven rotation causing protonation of a Glu residue at the cytoplasmic half-channel and subsequent deprotonation of a Glu residue at a luminal half-channel. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_8363.map.gz | 28.4 MB | EMDB map data format | |
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Header (meta data) | emd-8363-v30.xml emd-8363.xml | 11.7 KB 11.7 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_8363_fsc.xml | 7 KB | Display | FSC data file |
Images | emd_8363.png | 107.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-8363 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-8363 | HTTPS FTP |
-Validation report
Summary document | emd_8363_validation.pdf.gz | 78.4 KB | Display | EMDB validaton report |
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Full document | emd_8363_full_validation.pdf.gz | 77.5 KB | Display | |
Data in XML | emd_8363_validation.xml.gz | 494 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8363 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8363 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_8363.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | The Cryo-EM map of yeast Vo lacking the d subunit | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.554 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Vo region
Entire | Name: Vo region |
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Components |
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-Supramolecule #1: Vo region
Supramolecule | Name: Vo region / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all Details: Vo region of Saccharomyces cerevisiae Vacuolar-type ATPase lacking the d subunit. |
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-Macromolecule #1: Vo
Macromolecule | Name: Vo / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Sequence | String: Multisuque nce |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 2.5 mg/mL |
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Buffer | pH: 7.5 |
Grid | Model: Maxtaform / Material: COPPER/RHODIUM / Support film - topology: HOLEY / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK III |
Details | Vo solubilized in Amphipol A8-35 (Anatrace) Buffer: 50 mM Tris-HCl pH 7.5, 150 mM NaCl |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Details | 70 micro meter objective aperture, illuminated area of 1.58 micro meter diameter |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Number real images: 4365 / Average exposure time: 21.0 sec. / Average electron dose: 100.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated defocus max: 3.4 µm / Calibrated defocus min: 0.8 µm / Calibrated magnification: 64350 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal magnification: 37000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |