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- EMDB-8725: Yeast V-ATPase in complex with Legionella pneumophila effector Si... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-8725 | ||||||||||||||||||
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Title | Yeast V-ATPase in complex with Legionella pneumophila effector SidK (rotational state 2) | ||||||||||||||||||
![]() | V-ATPase:SidK complex, rotational state 2 | ||||||||||||||||||
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![]() | V-ATPase / SidK / rotational state 2 / HYDROLASE | ||||||||||||||||||
Function / homology | ![]() vacuole-mitochondrion membrane contact site / cell wall mannoprotein biosynthetic process / ATPase-coupled ion transmembrane transporter activity / protein localization to vacuolar membrane / 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 / cell wall mannoprotein biosynthetic process / ATPase-coupled ion transmembrane transporter activity / protein localization to vacuolar membrane / 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 proton-transporting V-type ATPase, V1 domain / vacuolar transport / protein targeting to vacuole / vacuole organization / proton-transporting V-type ATPase complex / protein metabolic process / fungal-type vacuole / intein-mediated protein splicing / intron homing / cellular hyperosmotic response / vacuolar proton-transporting V-type ATPase complex / phosphatidylinositol-3,5-bisphosphate binding / vacuolar acidification / fungal-type vacuole membrane / proton transmembrane transporter activity / intracellular copper ion homeostasis / ATP metabolic process / H+-transporting two-sector ATPase / Neutrophil degranulation / proton-transporting ATPase activity, rotational mechanism / proton transmembrane transport / proton-transporting ATP synthase activity, rotational mechanism / 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 | ![]() ![]() ![]() | ||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 7.9 Å | ||||||||||||||||||
![]() | Zhao J | ||||||||||||||||||
Funding support | ![]() ![]()
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![]() | ![]() 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
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 59.6 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 32.9 KB 32.9 KB | Display Display | ![]() |
Images | ![]() | 62.4 KB | ||
Filedesc metadata | ![]() | 9.3 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 5voyMC ![]() 8724C ![]() 8726C ![]() 5uf5C ![]() 5ufkC ![]() 5voxC ![]() 5vozC C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | V-ATPase:SidK complex, rotational state 2 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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
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Sample components
+Entire : V-ATPase:SidK complex
+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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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE-PROPANE |
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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: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | ![]() Model: Tecnai F20 / Image courtesy: FEI Company |
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Image processing
Startup model | Type of model: EMDB MAP |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 7.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 11125 |
Initial angle assignment | Type: PROJECTION MATCHING |
Final angle assignment | Type: PROJECTION MATCHING |