+Open data
-Basic information
Entry | Database: PDB / ID: 8ojj | ||||||
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Title | Cryo-EM structure of the DnaD-NTD tetramer | ||||||
Components | DNA replication protein DnaD | ||||||
Keywords | DNA BINDING PROTEIN / Replication helicase loading / small cryo-EM structure | ||||||
Function / homology | Function and homology information | ||||||
Biological species | Bacillus subtilis subsp. subtilis str. 168 (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 5.47 Å | ||||||
Authors | Winterhalter, C. / Pelliciari, S. / Cronin, N. / Costa, T.R.D. / Murray, H. / Ilangovan, A. | ||||||
Funding support | United Kingdom, 1items
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Citation | Journal: Nucleic Acids Res / Year: 2023 Title: The DNA replication initiation protein DnaD recognises a specific strand of the Bacillus subtilis chromosome origin. Authors: Charles Winterhalter / Simone Pelliciari / Daniel Stevens / Stepan Fenyk / Elie Marchand / Nora B Cronin / Panos Soultanas / Tiago R D Costa / Aravindan Ilangovan / Heath Murray / Abstract: Genome replication is a fundamental biological activity shared by all organisms. Chromosomal replication proceeds bidirectionally from origins, requiring the loading of two helicases, one for each ...Genome replication is a fundamental biological activity shared by all organisms. Chromosomal replication proceeds bidirectionally from origins, requiring the loading of two helicases, one for each replisome. However, the molecular mechanisms underpinning helicase loading at bacterial chromosome origins (oriC) are unclear. Here we investigated the essential DNA replication initiation protein DnaD in the model organism Bacillus subtilis. A set of DnaD residues required for ssDNA binding was identified, and photo-crosslinking revealed that this ssDNA binding region interacts preferentially with one strand of oriC. Biochemical and genetic data support the model that DnaD recognizes a new single-stranded DNA (ssDNA) motif located in oriC, the DnaD Recognition Element (DRE). Considered with single particle cryo-electron microscopy (cryo-EM) imaging of DnaD, we propose that the location of the DRE within oriC orchestrates strand-specific recruitment of helicase during DNA replication initiation. These findings significantly advance our mechanistic understanding of bidirectional replication from a bacterial chromosome origin. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8ojj.cif.gz | 107.5 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8ojj.ent.gz | 79.9 KB | Display | PDB format |
PDBx/mmJSON format | 8ojj.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8ojj_validation.pdf.gz | 1.1 MB | Display | wwPDB validaton report |
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Full document | 8ojj_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 8ojj_validation.xml.gz | 28 KB | Display | |
Data in CIF | 8ojj_validation.cif.gz | 38.9 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/oj/8ojj ftp://data.pdbj.org/pub/pdb/validation_reports/oj/8ojj | HTTPS FTP |
-Related structure data
Related structure data | 16914MC 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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-Components
#1: Protein | Mass: 27675.715 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Bacillus subtilis subsp. subtilis str. 168 (bacteria) Strain: 168 / Gene: dnaD, BSU22350 / Production host: Escherichia coli (E. coli) / References: UniProt: P39787 |
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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: Homo Tetrameric complex of DnaD N-terminal domain / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Source (natural) | Organism: Bacillus subtilis (bacteria) |
Source (recombinant) | Organism: Escherichia coli (E. coli) |
Buffer solution | pH: 7 |
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: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2700 nm / Nominal defocus min: 700 nm |
Image recording | Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
EM software | Name: EPU / Category: image acquisition |
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CTF correction | Type: NONE |
3D reconstruction | Resolution: 5.47 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 443529 / Symmetry type: POINT |
Atomic model building | Protocol: RIGID BODY FIT |