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- PDB-9iob: Cryo-EM structure of the hexameric DRT9-ncRNA complex -

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Basic information

Entry
Database: PDB / ID: 9iob
TitleCryo-EM structure of the hexameric DRT9-ncRNA complex
Components
  • RNA (177-MER)
  • RNA-dependent DNA polymerase
KeywordsANTIVIRAL PROTEIN/RNA / RNA BINDING / PROTEIN-RNA COMPLEX / ANTIVIRAL PROTEIN-RNA complex
Function / homology
Function and homology information


: / Reverse transcriptase domain / Reverse transcriptase (RNA-dependent DNA polymerase) / Reverse transcriptase (RT) catalytic domain profile. / DNA/RNA polymerase superfamily
Similarity search - Domain/homology
: / RNA / RNA (> 10) / RNA (> 100) / RNA-dependent DNA polymerase
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.62 Å
AuthorsZhang, J.T. / Song, X.Y. / Xia, Y.S. / Liu, Y.J. / Jia, N.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Science / Year: 2025
Title: Bacterial reverse transcriptase synthesizes long poly-A-rich cDNA for antiphage defense.
Authors: Xin-Yi Song / Yushan Xia / Jun-Tao Zhang / Yu-Jun Liu / Hua Qi / Xin-Yang Wei / Hailiang Hu / Yu Xia / Xue Liu / Ying-Fei Ma / Ning Jia /
Abstract: Prokaryotic defense-associated reverse transcriptases (DRTs) were recently identified with antiviral functions; however, their functional mechanisms remain largely unexplored. Here we show that DRT9 ...Prokaryotic defense-associated reverse transcriptases (DRTs) were recently identified with antiviral functions; however, their functional mechanisms remain largely unexplored. Here we show that DRT9 forms a hexameric complex with its upstream non-coding RNA (ncRNA) to mediate antiphage defense by inducing cell growth arrest via abortive infection. Upon phage infection, the phage-encoded ribonucleotide reductase NrdAB complex elevates intracellular dATP levels, activating DRT9 to synthesize long, poly-A-rich single-stranded cDNA, which likely sequesters the essential phage SSB protein and disrupts phage propagation. We further determined the cryo-electron microscopy structure of the DRT9-ncRNA hexamer complex, providing mechanistic insights into its cDNA synthesis. These findings highlight the diversity of RT-based antiviral defense mechanisms, expand our understanding of RT biological functions, and provide a structural basis for developing DRT9-based biotechnological tools.
History
DepositionJul 8, 2024Deposition site: PDBJ / Processing site: PDBC
Revision 1.0May 14, 2025Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: RNA-dependent DNA polymerase
B: RNA (177-MER)
C: RNA-dependent DNA polymerase
D: RNA (177-MER)
E: RNA-dependent DNA polymerase
F: RNA (177-MER)
G: RNA-dependent DNA polymerase
H: RNA (177-MER)
I: RNA-dependent DNA polymerase
J: RNA (177-MER)
K: RNA-dependent DNA polymerase
L: RNA (177-MER)


Theoretical massNumber of molelcules
Total (without water)690,67412
Polymers690,67412
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein
RNA-dependent DNA polymerase


Mass: 58318.961 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: GQA06_05700
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: A0A6D0I497
#2: RNA chain
RNA (177-MER)


Mass: 56793.293 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli)
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: GenBank: 2459894762
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: DRT9-ncRNA hexamer complex / Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Source (natural)Organism: Escherichia coli (E. coli)
Source (recombinant)Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1500 nm / Cs: 0.01 mm / C2 aperture diameter: 70 µm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.20.1_4487: / Category: model refinement
CTF correctionType: PHASE FLIPPING ONLY
3D reconstructionResolution: 2.62 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 220612 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00449812
ELECTRON MICROSCOPYf_angle_d0.64572300
ELECTRON MICROSCOPYf_dihedral_angle_d17.6815954
ELECTRON MICROSCOPYf_chiral_restr0.0388850
ELECTRON MICROSCOPYf_plane_restr0.0065274

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