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- EMDB-52583: Cryo-EM structure of retron Eco2 (Ec67) -

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Basic information

Entry
Database: EMDB / ID: EMD-52583
TitleCryo-EM structure of retron Eco2 (Ec67)
Map data
Sample
  • Complex: Ternary complex of retron Eco2 (Ec67) with RNA and DNA
    • DNA: msDNA (67-MER)
    • Protein or peptide: Retron Ec67 protein
    • RNA: RNA (132-MER)
KeywordsRetron / Reverse transcriptase / DNA / RNA / TOPRIM / RNaseH / msDNA / ANTIVIRAL PROTEIN
Function / homology
Function and homology information


ribonuclease H / RNA-directed DNA polymerase / RNA-directed DNA polymerase activity / RNA-DNA hybrid ribonuclease activity / defense response to virus / RNA binding / metal ion binding
Similarity search - Function
: / RNA-directed DNA polymerase (reverse transcriptase), msDNA / : / Reverse transcriptase (RNA-dependent DNA polymerase) / Reverse transcriptase domain / Reverse transcriptase (RT) catalytic domain profile. / DNA/RNA polymerase superfamily
Similarity search - Domain/homology
Biological speciesEscherichia coli NCTC 86 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsJasnauskaite M / Miksys A / Skorupskaite A / Malinauskaite L / Pausch P
Funding supportEuropean Union, 1 items
OrganizationGrant numberCountry
European Molecular Biology Organization (EMBO)5342-2023European Union
CitationJournal: Nat Struct Mol Biol / Year: 2026
Title: Structure and mechanism of antiphage retron Eco2.
Authors: M Jasnauskaitė / J Juozapaitis / T Liegutė / R Grigaitis / A Skorupskaitė / W Steinchen / A Mikšys / L Truncaitė / K Kazlauskaitė / M F Torres Jiménez / S Khochare / G Dudas / G Bange ...Authors: M Jasnauskaitė / J Juozapaitis / T Liegutė / R Grigaitis / A Skorupskaitė / W Steinchen / A Mikšys / L Truncaitė / K Kazlauskaitė / M F Torres Jiménez / S Khochare / G Dudas / G Bange / L Malinauskaitė / I Songailienė / P Pausch /
Abstract: Retrons are prokaryotic reverse transcriptase systems that produce multicopy single-stranded DNA (msDNA), yet the principles by which they mediate antiviral defense remain largely unresolved. Here we ...Retrons are prokaryotic reverse transcriptase systems that produce multicopy single-stranded DNA (msDNA), yet the principles by which they mediate antiviral defense remain largely unresolved. Here we investigate the mechanism of Escherichia coli Eco2, a minimal retron composed of a single reverse transcriptase-nuclease fusion protein. Cryogenic electron microscopy and hydrogen/deuterium exchange mass spectrometry reveal the structures and dynamics of a trimeric nucleoprotein complex assembled within a branched msDNA scaffold, which cages the TOPRIM nucleases. We show that the phage-encoded endonuclease DenB initiates msDNA degradation, thereby unblocking the nuclease active sites. Activated Eco2 cuts transfer RNAs, resulting in translational shutdown for antiphage defense. We further identify ribosomal protein S1 as a putative RNA chaperone that associates with the msDNA precursor. These findings provide insights into the molecular mechanisms of minimal retrons and establish a structural basis for engineering of Eco2.
History
DepositionJan 20, 2025-
Header (metadata) releaseFeb 4, 2026-
Map releaseFeb 4, 2026-
UpdateFeb 25, 2026-
Current statusFeb 25, 2026Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_52583.png

Downloads & links

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Map

FileDownload / File: emd_52583.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 360 pix.
= 396. Å		1.1 Å/pix.
x 360 pix.
= 396. Å		1.1 Å/pix.
x 360 pix.
= 396. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.1
Minimum - Maximum-0.25281474 - 0.7501366
Average (Standard dev.)0.0080136275 (±0.016562385)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 396.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: #1

Fileemd_52583_additional_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_52583_half_map_1.map
Projections & Slices
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Histogram

Histogram (log scale)

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Half map: #1

Fileemd_52583_half_map_2.map
Projections & Slices
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Histogram

Histogram (log scale)

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Sample components

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Entire : Ternary complex of retron Eco2 (Ec67) with RNA and DNA

EntireName: Ternary complex of retron Eco2 (Ec67) with RNA and DNA
Components
  • Complex: Ternary complex of retron Eco2 (Ec67) with RNA and DNA
    • DNA: msDNA (67-MER)
    • Protein or peptide: Retron Ec67 protein
    • RNA: RNA (132-MER)

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Supramolecule #1: Ternary complex of retron Eco2 (Ec67) with RNA and DNA

SupramoleculeName: Ternary complex of retron Eco2 (Ec67) with RNA and DNA
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli NCTC 86 (bacteria) / Strain: 23

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Macromolecule #1: msDNA (67-MER)

MacromoleculeName: msDNA (67-MER) / type: dna / ID: 1 / Number of copies: 3 / Classification: DNA
Source (natural)Organism: Escherichia coli NCTC 86 (bacteria) / Strain: 23
Molecular weightTheoretical: 20.522113 KDa
SequenceString: (DT)(DC)(DC)(DT)(DT)(DC)(DG)(DC)(DA)(DC) (DA)(DG)(DC)(DA)(DC)(DA)(DC)(DC)(DT)(DG) (DC)(DC)(DG)(DT)(DA)(DT)(DA)(DG)(DC) (DT)(DC)(DT)(DG)(DA)(DA)(DT)(DC)(DA)(DA) (DG) (DG)(DA)(DT)(DT)(DT)(DT) ...String:
(DT)(DC)(DC)(DT)(DT)(DC)(DG)(DC)(DA)(DC) (DA)(DG)(DC)(DA)(DC)(DA)(DC)(DC)(DT)(DG) (DC)(DC)(DG)(DT)(DA)(DT)(DA)(DG)(DC) (DT)(DC)(DT)(DG)(DA)(DA)(DT)(DC)(DA)(DA) (DG) (DG)(DA)(DT)(DT)(DT)(DT)(DA)(DG) (DG)(DG)(DA)(DG)(DG)(DC)(DG)(DA)(DT)(DT) (DC)(DC) (DT)(DC)(DC)(DT)(DG)(DC)(DC)

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Macromolecule #2: Retron Ec67 protein

MacromoleculeName: Retron Ec67 protein / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO / EC number: RNA-directed DNA polymerase
Source (natural)Organism: Escherichia coli NCTC 86 (bacteria) / Strain: 23
Molecular weightTheoretical: 68.511922 KDa
Recombinant expressionOrganism: Escherichia coli DH5[alpha] (bacteria)
SequenceString: MTKTSKLDAL RAATSREDLA KILDVKLVFL TNVLYRIGSD NQYTQFTIPK KGKGVRTISA PTDRLKDIQR RICDLLSDCR DEIFAIRKI SNNYSFGFER GKSIILNAYK HRGKQIILNI DLKDFFESFN FGRVRGYFLS NQDFLLNPVV ATTLAKAACY N GTLPQGSP ...String:
MTKTSKLDAL RAATSREDLA KILDVKLVFL TNVLYRIGSD NQYTQFTIPK KGKGVRTISA PTDRLKDIQR RICDLLSDCR DEIFAIRKI SNNYSFGFER GKSIILNAYK HRGKQIILNI DLKDFFESFN FGRVRGYFLS NQDFLLNPVV ATTLAKAACY N GTLPQGSP CSPIISNLIC NIMDMRLAKL AKKYGCTYSR YADDITISTN KNTFPLEMAT VQPEGVVLGK VLVKEIENSG FE INDSKTR LTYKTSRQEV TGLTVNRIVN IDRCYYKKTR ALAHALYRTG EYKVPDENGV LVSGGLDKLE GMFGFIDQVD KFN NIKKKL NKQPDRYVLT NATLHGFKLK LNAREKAYSK FIYYKFFHGN TCPTIITEGK TDRIYLKAAL HSLETSYPEL FREK TDSKK KEINLNIFKS NEKTKYFLDL SGGTADLKKF VERYKNNYAS YYGSVPKQPV IMVLDNDTGP SDLLNFLRNK VKSCP DDVT EMRKMKYIHV FYNLYIVLTP LSPSGEQTSM EDLFPKDILD IKIDGKKFNK NNDGDSKTEY GKHIFSMRVV RDKKRK IDF KAFCCIFDAI KDIKEHYKLM LNSGSWSHPQ FEK

UniProtKB: Retron Ec67 protein

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Macromolecule #3: RNA (132-MER)

MacromoleculeName: RNA (132-MER) / type: rna / ID: 3 / Number of copies: 3
Source (natural)Organism: Escherichia coli NCTC 86 (bacteria) / Strain: 23
Molecular weightTheoretical: 42.593203 KDa
SequenceString:
CACGCAUGUA GGCAGAUUUG UUGGUUGUGA AUCGCAACCA GUGGCCUUAA UGGCAGGAGG AAUCGCCUCC CUAAAAUCCU UGAUUCAGA GCUAUACGGC AGGUGUGCUG UGCGAAGGAG UGCCUGCAUG CGU

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration2 mg/mL
BufferpH: 8 / Component:
ConcentrationName
100.0 mMTris-HCl
150.0 mMNaCl
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 7 sec. / Pretreatment - Atmosphere: AIR / Details: 20 mA current
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS GLACIOS
SoftwareName: EPU (ver. 3.2)
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 1988 / Average exposure time: 46.33 sec. / Average electron dose: 30.31 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated defocus max: 2.0 µm / Calibrated defocus min: 1.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 92000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN

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Image processing

Particle selectionNumber selected: 544986
CTF correctionSoftware - Name: cryoSPARC (ver. 4.3) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL / In silico model: AlphaFold
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.3) / Number images used: 504525
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
Output model

PDB-9i2f:
Cryo-EM structure of retron Eco2 (Ec67)

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