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Yorodumi- EMDB-8724: Yeast V-ATPase in complex with Legionella pneumophila effector Si... -
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-Basic information
Entry | Database: EMDB / ID: EMD-8724 | ||||||||||||||||||
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Title | Yeast V-ATPase in complex with Legionella pneumophila effector SidK (rotational state 1) | ||||||||||||||||||
Map data | V-ATPase:SidK complex, rotational state 1 | ||||||||||||||||||
Sample | SidK != V-ATPase:SidK complex SidK
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Keywords | V-ATPase / SidK / rotational state 1 / HYDROLASE | ||||||||||||||||||
Function / homology | Function and homology information vacuole-mitochondrion membrane contact site / protein localization to vacuolar membrane / 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 ...vacuole-mitochondrion membrane contact site / protein localization to vacuolar membrane / 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 / proteasome storage granule assembly / P-type proton-exporting transporter activity / pexophagy / endosomal lumen acidification / vacuolar proton-transporting V-type ATPase, V0 domain / vacuolar transport / vacuolar proton-transporting V-type ATPase, V1 domain / protein targeting to vacuole / vacuole organization / cellular hyperosmotic response / proton-transporting V-type ATPase complex / vacuolar proton-transporting V-type ATPase complex / fungal-type vacuole / intein-mediated protein splicing / intron homing / vacuolar acidification / phosphatidylinositol-3,5-bisphosphate binding / fungal-type vacuole membrane / proton transmembrane transporter activity / intracellular copper ion homeostasis / H+-transporting two-sector ATPase / ATP metabolic process / Neutrophil degranulation / proton-transporting ATPase activity, rotational mechanism / proton-transporting ATP synthase activity, rotational mechanism / proton transmembrane transport / cell periphery / transmembrane transport / intracellular calcium ion homeostasis / cytoplasmic stress granule / endocytosis / ATPase binding / protein-containing complex assembly / endonuclease activity / intracellular iron ion homeostasis / Hydrolases; Acting on ester bonds / membrane raft / Golgi membrane / mRNA binding / ATP hydrolysis activity / DNA binding / ATP binding / membrane / cytoplasm Similarity search - Function | ||||||||||||||||||
Biological species | Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) / Legionella pneumophila subsp. pneumophila ATCC 43290 (bacteria) / Saccharomyces cerevisiae (brewer's yeast) | ||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 6.8 Å | ||||||||||||||||||
Authors | Zhao J | ||||||||||||||||||
Funding support | Canada, United States, 5 items
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Citation | Journal: PLoS Pathog / Year: 2017 Title: Molecular basis for the binding and modulation of V-ATPase by a bacterial effector protein. Authors: Jianhua Zhao / Ksenia Beyrakhova / Yao Liu / Claudia P Alvarez / Stephanie A Bueler / Li Xu / Caishuang Xu / Michal T Boniecki / Voula Kanelis / Zhao-Qing Luo / Miroslaw Cygler / John L Rubinstein / Abstract: Intracellular pathogenic bacteria evade the immune response by replicating within host cells. Legionella pneumophila, the causative agent of Legionnaires' Disease, makes use of numerous effector ...Intracellular pathogenic bacteria evade the immune response by replicating within host cells. Legionella pneumophila, the causative agent of Legionnaires' Disease, makes use of numerous effector proteins to construct a niche supportive of its replication within phagocytic cells. The L. pneumophila effector SidK was identified in a screen for proteins that reduce the activity of the proton pumping vacuolar-type ATPases (V-ATPases) when expressed in the yeast Saccharomyces cerevisae. SidK is secreted by L. pneumophila in the early stages of infection and by binding to and inhibiting the V-ATPase, SidK reduces phagosomal acidification and promotes survival of the bacterium inside macrophages. We determined crystal structures of the N-terminal region of SidK at 2.3 Å resolution and used single particle electron cryomicroscopy (cryo-EM) to determine structures of V-ATPase:SidK complexes at ~6.8 Å resolution. SidK is a flexible and elongated protein composed of an α-helical region that interacts with subunit A of the V-ATPase and a second region of unknown function that is flexibly-tethered to the first. SidK binds V-ATPase strongly by interacting via two α-helical bundles at its N terminus with subunit A. In vitro activity assays show that SidK does not inhibit the V-ATPase completely, but reduces its activity by ~40%, consistent with the partial V-ATPase deficiency phenotype its expression causes in yeast. The cryo-EM analysis shows that SidK reduces the flexibility of the A-subunit that is in the 'open' conformation. Fluorescence experiments indicate that SidK binding decreases the affinity of V-ATPase for a fluorescent analogue of ATP. Together, these results reveal the structural basis for the fine-tuning of V-ATPase activity by SidK. | ||||||||||||||||||
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_8724.map.gz | 59.6 MB | EMDB map data format | |
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Header (meta data) | emd-8724-v30.xml emd-8724.xml | 32.8 KB 32.8 KB | Display Display | EMDB header |
Images | emd_8724.png | 63.7 KB | ||
Filedesc metadata | emd-8724.cif.gz | 9.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-8724 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-8724 | HTTPS FTP |
-Validation report
Summary document | emd_8724_validation.pdf.gz | 576.6 KB | Display | EMDB validaton report |
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Full document | emd_8724_full_validation.pdf.gz | 576.2 KB | Display | |
Data in XML | emd_8724_validation.xml.gz | 6.3 KB | Display | |
Data in CIF | emd_8724_validation.cif.gz | 7.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8724 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8724 | HTTPS FTP |
-Related structure data
Related structure data | 5voxMC 8725C 8726C 5uf5C 5ufkC 5voyC 5vozC C: citing same article (ref.) M: atomic model generated by this map |
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Similar 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_8724.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | V-ATPase:SidK complex, rotational state 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.45 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 : SidK
+Supramolecule #1: V-ATPase:SidK complex
+Supramolecule #2: V-ATPase
+Supramolecule #3: SidK
+Macromolecule #1: V-type proton ATPase catalytic subunit A,V-type proton ATPase cat...
+Macromolecule #2: V-type proton ATPase subunit B
+Macromolecule #3: V-type proton ATPase subunit E
+Macromolecule #4: V-type proton ATPase subunit G
+Macromolecule #5: V-type proton ATPase subunit D
+Macromolecule #6: V-type proton ATPase subunit F
+Macromolecule #7: V-type proton ATPase subunit C
+Macromolecule #8: V-type proton ATPase subunit H
+Macromolecule #9: V-type proton ATPase subunit d
+Macromolecule #10: V-type proton ATPase subunit c''
+Macromolecule #11: V-type proton ATPase subunit c'
+Macromolecule #12: V-type proton ATPase subunit c
+Macromolecule #13: V-type proton ATPase subunit a, vacuolar isoform
+Macromolecule #14: V-type proton ATPase subunit e
+Macromolecule #15: V-type proton ATPase subunit f
+Macromolecule #16: effector protein SidK
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE-PROPANE |
-Electron microscopy
Microscope | FEI TECNAI F20 |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 1.2 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | Model: Tecnai F20 / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: EMDB MAP |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 6.8 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 23924 |
Initial angle assignment | Type: PROJECTION MATCHING |
Final angle assignment | Type: PROJECTION MATCHING |