+Open data
-Basic information
Entry | Database: PDB / ID: 6suf | ||||||
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Title | Structure of Photorhabdus luminescens Tc holotoxin pore | ||||||
Components |
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Keywords | TOXIN / Complex / Holotoxin / Photorhabdus / Insecticidal / Translocation | ||||||
Function / homology | Function and homology information | ||||||
Biological species | Photorhabdus luminescens (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||
Authors | Roderer, D. / Raunser, S. | ||||||
Funding support | Germany, 1items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2019 Title: Structure of a Tc holotoxin pore provides insights into the translocation mechanism. Authors: Daniel Roderer / Oliver Hofnagel / Roland Benz / Stefan Raunser / Abstract: Tc toxins are modular toxin systems of insect and human pathogenic bacteria. They are composed of a 1.4-MDa pentameric membrane translocator (TcA) and a 250-kDa cocoon (TcB and TcC) encapsulating the ...Tc toxins are modular toxin systems of insect and human pathogenic bacteria. They are composed of a 1.4-MDa pentameric membrane translocator (TcA) and a 250-kDa cocoon (TcB and TcC) encapsulating the 30-kDa toxic enzyme (C terminus of TcC). Binding of Tc toxins to target cells and a pH shift trigger the conformational transition from the soluble prepore state to the membrane-embedded pore. Subsequently, the toxic enzyme is translocated and released into the cytoplasm. A high-resolution structure of a holotoxin embedded in membranes is missing, leaving open the question of whether TcB-TcC has an influence on the conformational transition of TcA. Here we show in atomic detail a fully assembled 1.7-MDa Tc holotoxin complex from in the membrane. We find that the 5 TcA protomers conformationally adapt to fit around the cocoon during the prepore-to-pore transition. The architecture of the Tc toxin complex allows TcB-TcC to bind to an already membrane-embedded TcA pore to form a holotoxin. Importantly, assembly of the holotoxin at the membrane results in spontaneous translocation of the toxic enzyme, indicating that this process is not driven by a proton gradient or other energy source. Mammalian lipids with zwitterionic head groups are preferred over other lipids for the integration of Tc toxins. In a nontoxic Tc toxin variant, we can visualize part of the translocating toxic enzyme, which transiently interacts with alternating negative charges and hydrophobic stretches of the translocation channel, providing insights into the mechanism of action of Tc toxins. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6suf.cif.gz | 2.3 MB | Display | PDBx/mmCIF format |
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PDB format | pdb6suf.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 6suf.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6suf_validation.pdf.gz | 955 KB | Display | wwPDB validaton report |
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Full document | 6suf_full_validation.pdf.gz | 1 MB | Display | |
Data in XML | 6suf_validation.xml.gz | 307 KB | Display | |
Data in CIF | 6suf_validation.cif.gz | 482 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/su/6suf ftp://data.pdbj.org/pub/pdb/validation_reports/su/6suf | HTTPS FTP |
-Related structure data
Related structure data | 10313MC 6sueC 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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1 |
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-Components
#1: Protein | Mass: 283230.375 Da / Num. of mol.: 5 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Photorhabdus luminescens (bacteria) / Gene: tcdA, tcdA1 / Production host: Escherichia coli (E. coli) / References: UniProt: Q9RN43 #2: Protein | | Mass: 274188.250 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Photorhabdus luminescens (bacteria) / Gene: tcdB2, TccC3 / Production host: Escherichia coli (E. coli) / References: UniProt: Q8GF99, UniProt: Q8GF97 |
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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: Tc holotoxin complex in the pore form, formed by the TcdA1 pentamer and TcdB2-TccC3 Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Value: 1.7 MDa / Experimental value: NO |
Source (natural) | Organism: Photorhabdus luminescens (bacteria) |
Source (recombinant) | Organism: Escherichia coli (E. coli) |
Buffer solution | pH: 8 |
Specimen | Conc.: 0.1 mg/ml / 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: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 60 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 64806 / Symmetry type: POINT |