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- EMDB-48616: Structure of TIGR-TasR in complex with tigRNA and target DNA afte... -

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Basic information

Entry
Database: EMDB / ID: EMD-48616
TitleStructure of TIGR-TasR in complex with tigRNA and target DNA after DNA cleavage
Map dataSharpened map of TIGR-TasR in complex with tigRNA and target DNA after DNA cleavage
Sample
  • Complex: TIGR-TasR product complex
    • Protein or peptide: Transposase IS116/IS110/IS902 C-terminal domain-containing protein
    • RNA: tigRNA
  • DNA: Target DNA, spacer A targeted strand, 5' of cut
  • DNA: Target DNA, spacer A targeted strand, 3' of cut
  • DNA: Target DNA, spacer B targeted strand, 5' of cut
  • DNA: Target DNA, spacer B targeted strand, 3' of cut
  • Ligand: MAGNESIUM ION
Keywordsnuclease / RNA-guided / Nop domain / DNA-binding protein / DNA-RNA HYBRID / DNA BINDING PROTEIN-RNA-DNA complex
Function / homologyTransposase, IS116/IS110/IS902 / Transposase IS116/IS110/IS902 family / transposase activity / DNA transposition / DNA binding / Transposase IS116/IS110/IS902 C-terminal domain-containing protein
Function and homology information
Biological speciesThermoproteota archaeon (archaea) / Escherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.05 Å
AuthorsWilkinson ME / Zhang F
Funding support United States, 1 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Science / Year: 2025
Title: TIGR-Tas: A family of modular RNA-guided DNA-targeting systems in prokaryotes and their viruses.
Authors: Guilhem Faure / Makoto Saito / Max E Wilkinson / Natalia Quinones-Olvera / Peiyu Xu / Daniel Flam-Shepherd / Stephanie Kim / Nishith Reddy / Shiyou Zhu / Lilia Evgeniou / Eugene V Koonin / ...Authors: Guilhem Faure / Makoto Saito / Max E Wilkinson / Natalia Quinones-Olvera / Peiyu Xu / Daniel Flam-Shepherd / Stephanie Kim / Nishith Reddy / Shiyou Zhu / Lilia Evgeniou / Eugene V Koonin / Rhiannon K Macrae / Feng Zhang /
Abstract: RNA-guided systems provide remarkable versatility, enabling diverse biological functions. Through iterative structural and sequence homology-based mining starting with a guide RNA-interaction domain ...RNA-guided systems provide remarkable versatility, enabling diverse biological functions. Through iterative structural and sequence homology-based mining starting with a guide RNA-interaction domain of Cas9, we identified a family of RNA-guided DNA-targeting proteins in phage and parasitic bacteria. Each system consists of a tandem interspaced guide RNA (TIGR) array and a TIGR-associated (Tas) protein containing a nucleolar protein (Nop) domain, sometimes fused to HNH (TasH)- or RuvC (TasR)-nuclease domains. We show that TIGR arrays are processed into 36-nucleotide RNAs (tigRNAs) that direct sequence-specific DNA binding through a tandem-spacer targeting mechanism. TasR can be reprogrammed for precise DNA cleavage, including in human cells. The structure of TasR reveals striking similarities to box C/D small nucleolar ribonucleoproteins and IS110 RNA-guided transposases, providing insights into the evolution of diverse RNA-guided systems.
History
DepositionJan 12, 2025-
Header (metadata) releaseMar 5, 2025-
Map releaseMar 5, 2025-
UpdateMay 14, 2025-
Current statusMay 14, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_48616.png

Downloads & links

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Map

FileDownload / File: emd_48616.map.gz / Format: CCP4 / Size: 27 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened map of TIGR-TasR in complex with tigRNA and target DNA after DNA cleavage
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.98 Å/pix.
x 192 pix.
= 189.091 Å		0.98 Å/pix.
x 192 pix.
= 189.091 Å		0.98 Å/pix.
x 192 pix.
= 189.091 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.98485 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.025
Minimum - Maximum-0.09386886 - 0.13973059
Average (Standard dev.)0.000017526101 (±0.0047891247)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions192192192
Spacing192192192
CellA=B=C: 189.0912 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_48616_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: TIGR-TasR product complex refinement, half-map 1

Fileemd_48616_half_map_1.map
AnnotationTIGR-TasR product complex refinement, half-map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: TIGR-TasR product complex refinement, half-map 2

Fileemd_48616_half_map_2.map
AnnotationTIGR-TasR product complex refinement, half-map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : TIGR-TasR product complex

EntireName: TIGR-TasR product complex
Components
  • Complex: TIGR-TasR product complex
    • Protein or peptide: Transposase IS116/IS110/IS902 C-terminal domain-containing protein
    • RNA: tigRNA
  • DNA: Target DNA, spacer A targeted strand, 5' of cut
  • DNA: Target DNA, spacer A targeted strand, 3' of cut
  • DNA: Target DNA, spacer B targeted strand, 5' of cut
  • DNA: Target DNA, spacer B targeted strand, 3' of cut
  • Ligand: MAGNESIUM ION

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Supramolecule #1: TIGR-TasR product complex

SupramoleculeName: TIGR-TasR product complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Thermoproteota archaeon (archaea)
Molecular weightTheoretical: 130 KDa

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Macromolecule #1: Transposase IS116/IS110/IS902 C-terminal domain-containing protein

MacromoleculeName: Transposase IS116/IS110/IS902 C-terminal domain-containing protein
type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Thermoproteota archaeon (archaea)
Molecular weightTheoretical: 38.276855 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MDNDITILAV DWSHEERKLA IFDGKKIRKK LPEPSSDVII VAENIPQKYA APFIEVGAKV LRCSTNATAD ARKNYQKKVD AAFAKNDEN DSKVIWALYQ THPELFREMK LEPPLSSYYA IFKDYQEVRI RTGNRLYSDR TDAMEEFFKI VKKGEHELKK A VDKELENH ...String:
MDNDITILAV DWSHEERKLA IFDGKKIRKK LPEPSSDVII VAENIPQKYA APFIEVGAKV LRCSTNATAD ARKNYQKKVD AAFAKNDEN DSKVIWALYQ THPELFREMK LEPPLSSYYA IFKDYQEVRI RTGNRLYSDR TDAMEEFFKI VKKGEHELKK A VDKELENH PVYTQWLQHI KGIGPVVAGG LISLIGDIDR FDSVSKLWAY AGYSVDNGKV QKRKKGVASN WKNKIRTHCY NI VDSFIKQ RTSVYRELYD AEKARQRPKV ESDGHAHNRA VRKVAKVFLQ HYWVVSRELA GFSVSKPWIL EHGGHVDYIK PPH WNKVEI KPCKP

UniProtKB: Transposase IS116/IS110/IS902 C-terminal domain-containing protein

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Macromolecule #2: tigRNA

MacromoleculeName: tigRNA / type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: Thermoproteota archaeon (archaea)
Molecular weightTheoretical: 11.484882 KDa
SequenceString:
AGCCAUGCAA GCCCUGAAAC CCAUUGCCUU AGUGCG

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Macromolecule #3: Target DNA, spacer A targeted strand, 5' of cut

MacromoleculeName: Target DNA, spacer A targeted strand, 5' of cut / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 10.537799 KDa
SequenceString:
(DT)(DC)(DA)(DG)(DC)(DG)(DT)(DT)(DA)(DA) (DA)(DA)(DA)(DA)(DT)(DT)(DG)(DA)(DT)(DG) (DA)(DT)(DT)(DG)(DC)(DC)(DT)(DT)(DA) (DG)(DG)(DG)(DG)(DC)

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Macromolecule #4: Target DNA, spacer A targeted strand, 3' of cut

MacromoleculeName: Target DNA, spacer A targeted strand, 3' of cut / type: dna / ID: 4 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 7.999197 KDa
SequenceString:
(DT)(DT)(DG)(DC)(DA)(DT)(DC)(DG)(DA)(DA) (DA)(DC)(DC)(DA)(DC)(DA)(DG)(DC)(DC)(DA) (DA)(DG)(DG)(DG)(DA)(DA)

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Macromolecule #5: Target DNA, spacer B targeted strand, 5' of cut

MacromoleculeName: Target DNA, spacer B targeted strand, 5' of cut / type: dna / ID: 5 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 10.392662 KDa
SequenceString:
(DT)(DT)(DC)(DC)(DC)(DT)(DT)(DG)(DG)(DC) (DT)(DG)(DT)(DG)(DG)(DT)(DT)(DT)(DC)(DG) (DA)(DT)(DG)(DC)(DA)(DA)(DG)(DC)(DC) (DC)(DC)(DT)(DA)(DA)

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Macromolecule #6: Target DNA, spacer B targeted strand, 3' of cut

MacromoleculeName: Target DNA, spacer B targeted strand, 3' of cut / type: dna / ID: 6 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 7.961164 KDa
SequenceString:
(DG)(DG)(DC)(DA)(DA)(DT)(DC)(DA)(DT)(DC) (DA)(DA)(DT)(DT)(DT)(DT)(DT)(DT)(DA)(DA) (DC)(DG)(DC)(DT)(DG)(DA)

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Macromolecule #7: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 7 / Number of copies: 4 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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

Concentration1 mg/mL
BufferpH: 7.9
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 14257 / Average exposure time: 1.09 sec. / Average electron dose: 45.4 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 130000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 10065225
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.05 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 5.0) / Number images used: 763089
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-9mty:
Structure of TIGR-TasR in complex with tigRNA and target DNA after DNA cleavage

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