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- PDB-9p4j: The structure of Retron Eco8 in Apo state -

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Basic information

Entry
Database: PDB / ID: 9p4j
TitleThe structure of Retron Eco8 in Apo state
Components
  • Retron Eco8 OLD nuclease
  • Retron Eco8 reverse transcriptase
  • msdDNA
  • msrRNA
KeywordsTransferase/Hydrolase/DNA/RNA / anti-phage / bacterial immunity / retron / DNA / RNA / reverse transcription / Transferase-Hydrolase-DNA-RNA complex
Function / homology
Function and homology information


nuclease activity / RNA-directed DNA polymerase / RNA-directed DNA polymerase activity / defense response to virus / Hydrolases; Acting on ester bonds / ATP hydrolysis activity / RNA binding / ATP binding / metal ion binding
Similarity search - Function
AAA domain, putative AbiEii toxin, Type IV TA system / : / RNA-directed DNA polymerase (reverse transcriptase), msDNA / : / ATPase, AAA-type, core / Reverse transcriptase domain / Reverse transcriptase (RNA-dependent DNA polymerase) / Reverse transcriptase (RT) catalytic domain profile. / DNA/RNA polymerase superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
: / DNA / DNA (> 10) / RNA / RNA (> 10) / Retron Eco8 OLD nuclease / Retron Eco8 reverse transcriptase
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.68 Å
AuthorsYu, C. / Wang, C. / Fu, T.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS) United States
CitationJournal: Mol Cell / Year: 2025
Title: Phage SSB detection by retron Eco8 msDNA unleashes nuclease-mediated immunity.
Authors: Chengzhi Yu / Chen Wang / Taylor Forman / Jiale Xie / Samantha Major / Matthew X Fang / Jewel E Voyer / Joe Pogliano / Tian-Min Fu /
Abstract: Retrons are antiphage defense systems that synthesize multicopy single-stranded DNA (msDNA) and are being adapted for genome engineering. The Escherichia coli retron Eco8 system comprises a reverse ...Retrons are antiphage defense systems that synthesize multicopy single-stranded DNA (msDNA) and are being adapted for genome engineering. The Escherichia coli retron Eco8 system comprises a reverse transcriptase (RT), an msDNA, and an overcoming lysogenization defect (OLD)-family nuclease (Eco8-OLD), reminiscent of Gabija. Here, we present the cryo-electron microscopy structure of the Eco8 supramolecular complex, a symmetric 4:4:4 assembly of RT, Eco8-OLD, and msDNA (a hybrid of msrRNA and msdDNA). The msDNA anchors RT and Eco8-OLD into a compact architecture that traps Eco8-OLD in an autoinhibited conformation. Upon phage infection, phage single-stranded DNA-binding proteins (SSBs) bind msdDNA, inducing conformational rearrangements that relieve Eco8-OLD autoinhibition and activate its non-specific DNA nuclease activity. This structural transition enables Eco8 to mount a robust antiphage response by restricting phage genome amplification. Our findings reveal a previously unrecognized mechanism of retron activation and highlight the central role of msDNA as a molecular switch in controlling retron-mediated antiphage defense.
History
DepositionJun 16, 2025Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 12, 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: Retron Eco8 OLD nuclease
B: Retron Eco8 OLD nuclease
C: Retron Eco8 OLD nuclease
D: Retron Eco8 OLD nuclease
E: Retron Eco8 reverse transcriptase
F: Retron Eco8 reverse transcriptase
G: Retron Eco8 reverse transcriptase
H: Retron Eco8 reverse transcriptase
I: msrRNA
M: msdDNA
J: msrRNA
K: msrRNA
L: msrRNA
N: msdDNA
O: msdDNA
P: msdDNA


Theoretical massNumber of molelcules
Total (without water)722,80116
Polymers722,80116
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
Retron Eco8 OLD nuclease


Mass: 87373.789 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: old, ERS139198_01420, Ga0119705_103344 / Production host: Escherichia coli (E. coli)
References: UniProt: P0DV58, Hydrolases; Acting on ester bonds
#2: Protein
Retron Eco8 reverse transcriptase / RT


Mass: 44213.145 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: ret, ERS139198_01421, Ga0119705_103345 / Production host: Escherichia coli (E. coli) / References: UniProt: P0DV59, RNA-directed DNA polymerase
#3: RNA chain
msrRNA


Mass: 25986.383 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Production host: Escherichia coli (E. coli) / References: GenBank: 1881611662
#4: DNA chain
msdDNA


Mass: 23126.848 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Production host: Escherichia coli (E. coli) / References: GenBank: 1881611662
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: The structure of Retron Eco8 complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Escherichia coli (E. coli)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE / Humidity: 100 %

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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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameVersionCategory
1cryoSPARCparticle selection
2PHENIX1.21_5207:model refinement
13cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING ONLY
3D reconstructionResolution: 2.68 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 421879 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00249521
ELECTRON MICROSCOPYf_angle_d0.43969452
ELECTRON MICROSCOPYf_dihedral_angle_d18.8220666
ELECTRON MICROSCOPYf_chiral_restr0.0368035
ELECTRON MICROSCOPYf_plane_restr0.0036756

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