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Yorodumi- PDB-5voy: Yeast V-ATPase in complex with Legionella pneumophila effector Si... -
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-Basic information
Entry | Database: PDB / ID: 5voy | ||||||||||||||||||
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Title | Yeast V-ATPase in complex with Legionella pneumophila effector SidK (rotational state 2) | ||||||||||||||||||
Components |
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Keywords | HYDROLASE / V-ATPase / SidK / rotational state 2 | ||||||||||||||||||
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 | Legionella pneumophila subsp. pneumophila ATCC 43290 (bacteria) Saccharomyces cerevisiae (brewer's yeast) | ||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 7.9 Å | ||||||||||||||||||
Authors | Zhao, J. | ||||||||||||||||||
Funding support | Canada, United States, 5items
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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 |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
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PDBx/mmCIF format | 5voy.cif.gz | 1.1 MB | Display | PDBx/mmCIF format |
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PDB format | pdb5voy.ent.gz | 708.5 KB | Display | PDB format |
PDBx/mmJSON format | 5voy.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 5voy_validation.pdf.gz | 1.1 MB | Display | wwPDB validaton report |
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Full document | 5voy_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 5voy_validation.xml.gz | 147.9 KB | Display | |
Data in CIF | 5voy_validation.cif.gz | 264.1 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/vo/5voy ftp://data.pdbj.org/pub/pdb/validation_reports/vo/5voy | HTTPS FTP |
-Related structure data
Related structure data | 8725MC 8724C 8726C 5uf5C 5ufkC 5voxC 5vozC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
-Protein , 2 types, 6 molecules ACEefg
#1: Protein | Mass: 67796.508 Da / Num. of mol.: 3 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c References: UniProt: P17255, H+-transporting two-sector ATPase, Hydrolases; Acting on ester bonds #16: Protein | Mass: 65490.320 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Legionella pneumophila subsp. pneumophila ATCC 43290 (bacteria) Gene: lp12_0990 / Production host: Escherichia coli (E. coli) / References: UniProt: G8UUS6 |
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-V-type proton ATPase subunit ... , 14 types, 27 molecules BDFGIKHJLMNOPQRSTUVWXYZabcd
#2: Protein | Mass: 57815.023 Da / Num. of mol.: 3 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P16140 #3: Protein | Mass: 26508.393 Da / Num. of mol.: 3 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P22203 #4: Protein | Mass: 12738.706 Da / Num. of mol.: 3 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P48836 #5: Protein | | Mass: 29235.023 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P32610 #6: Protein | | Mass: 13479.170 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P39111 #7: Protein | | Mass: 44241.352 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P31412 #8: Protein | | Mass: 54482.609 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P41807 #9: Protein | | Mass: 39822.484 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P32366 #10: Protein | | Mass: 22610.641 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P23968 #11: Protein | | Mass: 17046.361 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P32842 #12: Protein | Mass: 16357.501 Da / Num. of mol.: 8 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P25515 #13: Protein | | Mass: 95625.484 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P32563 #14: Protein | | Mass: 8387.065 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: Q3E7B6 #15: Protein | | Mass: 4613.678 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component |
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Source (natural) |
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Source (recombinant) | Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c | ||||||||||||||||||||||||
Buffer solution | pH: 7.5 | ||||||||||||||||||||||||
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||
Vitrification | Cryogen name: ETHANE-PROPANE |
-Electron microscopy imaging
Experimental equipment | Model: Tecnai F20 / Image courtesy: FEI Company |
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Microscopy | Model: FEI TECNAI F20 |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 1.2 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
Software | Name: PHENIX / Version: 1.11.1_2575: / Classification: refinement | ||||||||||||||||||||||||
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 7.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 11125 / Symmetry type: POINT | ||||||||||||||||||||||||
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