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Yorodumi- PDB-8qhc: Cryo-EM structure of SidH from Legionella pneumophila in complex ... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8qhc | ||||||
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Title | Cryo-EM structure of SidH from Legionella pneumophila in complex with LubX | ||||||
Components |
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Keywords | RNA BINDING PROTEIN / SidH / Legionella / tRNA-binding / EF-Tu-binding | ||||||
Function / homology | Function and homology information guanosine tetraphosphate binding / translation elongation factor activity / ubiquitin-protein transferase activity / protein ubiquitination / GTPase activity / GTP binding / cytoplasm Similarity search - Function | ||||||
Biological species | Legionella pneumophila (bacteria) Escherichia coli 'BL21-GoldpLysS AG' | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å | ||||||
Authors | Sharma, R. / Adams, M. / Bhogaraju, S. | ||||||
Funding support | France, 1items
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Citation | Journal: Nat Commun / Year: 2023 Title: Structural basis for the toxicity of Legionella pneumophila effector SidH. Authors: Rahul Sharma / Michael Adams / Simonne Griffith-Jones / Tobias Sahr / Laura Gomez-Valero / Felix Weis / Michael Hons / Sarah Gharbi / Rayene Berkane / Alexandra Stolz / Carmen Buchrieser / Sagar Bhogaraju / Abstract: Legionella pneumophila (LP) secretes more than 300 effectors into the host cytosol to facilitate intracellular replication. One of these effectors, SidH, 253 kDa in size with no sequence similarity ...Legionella pneumophila (LP) secretes more than 300 effectors into the host cytosol to facilitate intracellular replication. One of these effectors, SidH, 253 kDa in size with no sequence similarity to proteins of known function is toxic when overexpressed in host cells. SidH is regulated by the LP metaeffector LubX which targets SidH for degradation in a temporal manner during LP infection. The mechanism underlying the toxicity of SidH and its role in LP infection are unknown. Here, we determined the cryo-EM structure of SidH at 2.7 Å revealing a unique alpha helical arrangement with no overall similarity to known protein structures. Surprisingly, purified SidH came bound to a E. coli EF-Tu/t-RNA/GTP ternary complex which could be modeled into the cryo-EM density. Mutation of residues disrupting the SidH-tRNA interface and SidH-EF-Tu interface abolish the toxicity of overexpressed SidH in human cells, a phenotype confirmed in infection of Acanthamoeba castellani. We also present the cryo-EM structure of SidH in complex with a U-box domain containing ubiquitin ligase LubX delineating the mechanism of regulation of SidH. Our data provide the basis for the toxicity of SidH and into its regulation by the metaeffector LubX. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8qhc.cif.gz | 502.2 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8qhc.ent.gz | 307.1 KB | Display | PDB format |
PDBx/mmJSON format | 8qhc.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/qh/8qhc ftp://data.pdbj.org/pub/pdb/validation_reports/qh/8qhc | HTTPS FTP |
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-Related structure data
Related structure data | 18407MC 8qfsC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Protein , 3 types, 3 molecules CAD
#1: Protein | Mass: 43370.492 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: E2QJ06 |
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#3: Protein | Mass: 255113.719 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Legionella pneumophila (bacteria) / Gene: C3927_14265 Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: Q6RCQ4 |
#4: Protein | Mass: 30506.830 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Legionella pneumophila (bacteria) / Gene: C3927_14270 / Production host: Escherichia coli (E. coli) / References: UniProt: A0A2S6F0E3 |
-RNA chain , 1 types, 1 molecules B
#2: RNA chain | Mass: 24620.766 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
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-Non-polymers , 2 types, 2 molecules
#5: Chemical | ChemComp-MG / |
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#6: Chemical | ChemComp-GTP / |
-Details
Has ligand of interest | N |
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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 | Name: Quaternary complex of SidH with E.coli tRNA, EF-Tu and LubX Type: COMPLEX / Entity ID: #1-#4 / Source: RECOMBINANT |
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Source (natural) | Organism: Legionella pneumophila (bacteria) |
Source (recombinant) | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: TFS KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2200 nm / Nominal defocus min: 800 nm |
Image recording | Electron dose: 43.37 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION |
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3D reconstruction | Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 255829 / Symmetry type: POINT |