+Open data
-Basic information
Entry | Database: PDB / ID: 9ezx | ||||||
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Title | Vibrio cholerae DdmD apo complex | ||||||
Components | Helicase/UvrB N-terminal domain-containing protein | ||||||
Keywords | IMMUNE SYSTEM / Helicase / Nuclease / Complex / Effector | ||||||
Function / homology | Helicase/UvrB N-terminal domain-containing protein Function and homology information | ||||||
Biological species | Vibrio cholerae (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.55 Å | ||||||
Authors | Loeff, L. / Jinek, M. | ||||||
Funding support | European Union, 1items
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Citation | Journal: Science / Year: 2024 Title: Molecular mechanism of plasmid elimination by the DdmDE defense system. Authors: Luuk Loeff / David W Adams / Christelle Chanez / Sandrine Stutzmann / Laurie Righi / Melanie Blokesch / Martin Jinek / Abstract: Seventh-pandemic strains contain two pathogenicity islands that encode the DNA defense modules DdmABC and DdmDE. In this study, we used cryogenic electron microscopy to determine the mechanistic ...Seventh-pandemic strains contain two pathogenicity islands that encode the DNA defense modules DdmABC and DdmDE. In this study, we used cryogenic electron microscopy to determine the mechanistic basis for plasmid defense by DdmDE. The helicase-nuclease DdmD adopts an autoinhibited dimeric architecture. The prokaryotic Argonaute protein DdmE uses a DNA guide to target plasmid DNA. The structure of the DdmDE complex, validated by in vivo mutational studies, shows that DNA binding by DdmE triggers disassembly of the DdmD dimer and loading of monomeric DdmD onto the nontarget DNA strand. In vitro studies indicate that DdmD translocates in the 5'-to-3' direction, while partially degrading the plasmid DNA. These findings provide critical insights into the mechanism of DdmDE systems in plasmid elimination. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 9ezx.cif.gz | 461.6 KB | Display | PDBx/mmCIF format |
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PDB format | pdb9ezx.ent.gz | 372.3 KB | Display | PDB format |
PDBx/mmJSON format | 9ezx.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 9ezx_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 9ezx_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 9ezx_validation.xml.gz | 80.5 KB | Display | |
Data in CIF | 9ezx_validation.cif.gz | 119.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ez/9ezx ftp://data.pdbj.org/pub/pdb/validation_reports/ez/9ezx | HTTPS FTP |
-Related structure data
Related structure data | 50090MC 9ezyC M: map data used to model this data C: citing same article (ref.) |
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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
#1: Protein | Mass: 136427.594 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Vibrio cholerae (bacteria) / Gene: VC_1771 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9KR72 |
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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: Dimeric complex of the DdmD protein / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT | |||||||||||||||
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Molecular weight | Value: 0.266 MDa / Experimental value: YES | |||||||||||||||
Source (natural) | Organism: Vibrio cholerae (bacteria) | |||||||||||||||
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) | |||||||||||||||
Buffer solution | pH: 8 | |||||||||||||||
Buffer component |
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Specimen | Conc.: 1.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: This sample was monodisperse. | |||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / 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 FIELD / Calibrated magnification: 130000 X / Nominal defocus max: 2400 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 59.98 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 2023352 | ||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.55 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 184227 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||
Atomic model building | Details: Initial model was generated with alpha fold / Source name: AlphaFold / Type: in silico model |