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- EMDB-73998: 1-methyl-pseudouridine L-21 ScaI Tetrahymena Ribozyme - extended ... -

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Basic information

Entry
Database: EMDB / ID: EMD-73998
Title1-methyl-pseudouridine L-21 ScaI Tetrahymena Ribozyme - extended conformation
Map dataUnsharpened full map
Sample
  • Complex: L21-ScaI Tetrahymena ribozyme transcribed with 1-methyl-pseudouridine
    • RNA: 1-methyl-pseudouridine L-21 ScaI Tetrahymena Ribozyme
KeywordsRNA / Tetrahymena ribozyme / m1Y / 1-methyl-pseudouridine / modified base
Biological speciesTetrahymena thermophila (eukaryote)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.5 Å
AuthorsMcRae EKS / Yang H
Funding support United States, 1 items
OrganizationGrant numberCountry
Cancer Prevention and Research Institute of Texas (CPRIT)RR230015 United States
CitationJournal: Nat Commun / Year: 2026
Title: Base modifications shift tertiary structure and activity in synthetic RNA origami and a natural ribozyme.
Authors: Deepak Kumar Yadav / Haoyun Yang / Sukyeong Lee / Ewan K S McRae /
Abstract: Modified nucleotide bases like 5-methylcytosine (m5C) and N1-methyl-pseudouridine (m1Ψ) are widely used to enhance stability and reduce immunogenicity in therapeutic RNAs, yet their impact on RNA ...Modified nucleotide bases like 5-methylcytosine (m5C) and N1-methyl-pseudouridine (m1Ψ) are widely used to enhance stability and reduce immunogenicity in therapeutic RNAs, yet their impact on RNA tertiary structure remains unclear. Here we investigate how these modifications influence folding and function in both a synthetic RNA origami nanostructure and the natural Tetrahymena ribozyme. Using cryo-EM, FRET, and biochemical assays, we find that modified bases impede proper maturation of RNA origami by stabilizing alternative coaxial stacking at key junctions, leading to dimerization. In the ribozyme, modifications shift the equilibrium between open and closed conformations, altering catalytic activity in a temperature-dependent manner. These effects arise primarily from changes in base-stacking energetics rather than base pairing. Our findings reveal that base modifications reshape RNA folding landscapes and structure-function relationships, underscoring the need to consider structural consequences when designing modified RNAs for synthetic biology and therapeutic applications.
History
DepositionNov 21, 2025-
Header (metadata) releaseDec 3, 2025-
Map releaseDec 3, 2025-
UpdateJun 17, 2026-
Current statusJun 17, 2026Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_73998.png

Downloads & links

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Map

FileDownload / File: emd_73998.map.gz / Format: CCP4 / Size: 16.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationUnsharpened full map
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
1.84 Å/pix.
x 162 pix.
= 298.08 Å		1.84 Å/pix.
x 162 pix.
= 298.08 Å		1.84 Å/pix.
x 162 pix.
= 298.08 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.84 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.04
Minimum - Maximum-0.053781256 - 0.34428602
Average (Standard dev.)0.0010709701 (±0.016212987)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions162162162
Spacing162162162
CellA=B=C: 298.08002 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: DeepEMhancer sharpened map

Fileemd_73998_additional_1.map
AnnotationDeepEMhancer sharpened map
Projections & Slices
AxesZYX

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Half map: Half map B

Fileemd_73998_half_map_1.map
AnnotationHalf map B
Projections & Slices
AxesZYX

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Density Histograms

Histogram

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Half map: Half map A

Fileemd_73998_half_map_2.map
AnnotationHalf map A
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AxesZYX

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Sample components

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Entire : L21-ScaI Tetrahymena ribozyme transcribed with 1-methyl-pseudouridine

EntireName: L21-ScaI Tetrahymena ribozyme transcribed with 1-methyl-pseudouridine
Components
  • Complex: L21-ScaI Tetrahymena ribozyme transcribed with 1-methyl-pseudouridine
    • RNA: 1-methyl-pseudouridine L-21 ScaI Tetrahymena Ribozyme

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Supramolecule #1: L21-ScaI Tetrahymena ribozyme transcribed with 1-methyl-pseudouridine

SupramoleculeName: L21-ScaI Tetrahymena ribozyme transcribed with 1-methyl-pseudouridine
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Tetrahymena thermophila (eukaryote)
Molecular weightTheoretical: 125 KDa

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Macromolecule #1: 1-methyl-pseudouridine L-21 ScaI Tetrahymena Ribozyme

MacromoleculeName: 1-methyl-pseudouridine L-21 ScaI Tetrahymena Ribozyme / type: rna / ID: 1 / Number of copies: 1
Source (natural)Organism: Tetrahymena thermophila (eukaryote)
Molecular weightTheoretical: 126.457586 KDa
SequenceString: GGAGGGAAAA G(B8H)(B8H)A(B8H)CAGGC A(B8H)GCACC(B8H)GG (B8H)AGC(B8H)AG(B8H)C(B8H) (B8H)(B8H)AAACCAA (B8H)AGA(B8H)(B8H)GCA(B8H) CGG(B8H)(B8H)(B8H)AAAA GGCAAGACCG (B8H) CAAA(B8H)(B8H)GC ...String:
GGAGGGAAAA G(B8H)(B8H)A(B8H)CAGGC A(B8H)GCACC(B8H)GG (B8H)AGC(B8H)AG(B8H)C(B8H) (B8H)(B8H)AAACCAA (B8H)AGA(B8H)(B8H)GCA(B8H) CGG(B8H)(B8H)(B8H)AAAA GGCAAGACCG (B8H) CAAA(B8H)(B8H)GC GGGAAAGGGG (B8H)CAACAGCCG (B8H)(B8H)CAG(B8H)ACCA AG(B8H)C(B8H)CAGGG GAAAC(B8H)(B8H)(B8H)G AGA(B8H)GGCC(B8H)(B8H) GCAAAGGG(B8H)A (B8H)GG(B8H)AA(B8H)AAG C (B8H)GACGGAC A(B8H)GG(B8H)CC(B8H)AA CCACGCAGCC AAG(B8H)CC(B8H)AAG (B8H)CAACAGA(B8H)C (B8H)(B8H)C(B8H)G(B8H)(B8H)GA (B8H)A(B8H)GGA(B8H)GCA G(B8H)(B8H)CACAGAC (B8H)AAA (B8H)G(B8H)CG G(B8H)CGGGGAAG A(B8H)G(B8H)A(B8H)(B8H)C(B8H)(B8H) C(B8H)CA(B8H)AAGA (B8H)A(B8H)AG(B8H)CGGA CC(B8H)C(B8H)CC(B8H)(B8H)A A(B8H)GGGAGC(B8H)A GCGGA(B8H)GAAG (B8H)GA(B8H)GCAAC AC(B8H)GGAGCCG C(B8H)GGGAAC(B8H)A A(B8H)(B8H)(B8H)G(B8H)A(B8H)GC G AAAG(B8H)A(B8H)A (B8H)(B8H)GA(B8H)(B8H)AG(B8H)(B8H) (B8H)(B8H)GGAG

GENBANK: GENBANK: X54512.1

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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
Details: SEC purified in 50mM HEPES pH 8.0, 50mM KCl, 5mM MgCl2
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE
DetailsSample was transcribed in vitro with 1-methyl-pseudouridine triphosphate in place of UTP. Purified by size exclusion chromatography and refolded by heat denaturation and cooling to room temperature prior to the addition of 10mM MgCl2

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Electron microscopy

MicroscopeTFS GLACIOS
Specialist opticsEnergy filter - Name: TFS Selectris X / Energy filter - Slit width: 10 eV
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 130000

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Image processing

Particle selectionNumber selected: 599317 / Details: Blob picking on denoised micrographs
CTF correctionSoftware - Name: cryoSPARC / Details: Patch CTF / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 7.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Software - details: non-uniform refinement / Number images used: 60417
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationNumber classes: 3 / Avg.num./class: 250000 / Software - Name: cryoSPARC
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

7xsn


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Target criteria: geometry minimization
Output model

PDB-9zbr:
1-methyl-pseudouridine L-21 ScaI Tetrahymena Ribozyme - extended conformation

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