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- EMDB-70040: N1Y-TC-6HBC type-2 dimer RNA -

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Basic information

Entry
Database: EMDB / ID: EMD-70040
TitleN1Y-TC-6HBC type-2 dimer RNA
Map data
Sample
  • Complex: Dimeric structure of a 6-helix bundle with a clasp helix transcribed with N1-methyl-pseudouridine.
    • RNA: Twist corrected 6HBC type 2 dimer transcribed with N1-methyl-pseudouridine
KeywordsRNA / origami / N1Y / N1-methyl-pseudouridine / modified base
Biological speciessynthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 12.0 Å
AuthorsMcRae EKS / Kumar DY
Funding support United States, 1 items
OrganizationGrant numberCountry
Cancer Prevention and Research Institute of Texas (CPRIT)RR230015 United States
CitationJournal: Res Sq / Year: 2025
Title: Base modifications reshape RNA folding landscapes and structure-function relationships in synthetic and natural RNAs.
Authors: Ewan McRae / Deepak Yadav / Haoyun Yang / Sukyeong Lee /
Abstract: Modified bases such as 5-methylcytosine (5mC) and N1-methyl-pseudouridine (N1Ψ) are widely used to reduce the immunogenicity of and enhance the stability and translational efficiency of therapeutic ...Modified bases such as 5-methylcytosine (5mC) and N1-methyl-pseudouridine (N1Ψ) are widely used to reduce the immunogenicity of and enhance the stability and translational efficiency of therapeutic RNAs, yet their effects on RNA 3D structure remain poorly understood. Here, we investigate how these base modifications influence the folding and function of structured RNAs using both an engineered RNA origami nanostructure and a naturally occurring ribozyme. We show that modified bases impair kinetic maturation of RNA nanostructures and promote alternative folding topologies via stabilization of noncanonical stacking arrangements. Cryo-EM and FRET experiments reveal that this conformational shift is driven not by changes in base pairing, but by altered stacking energetics at key junctions. Our findings highlight the need to consider structural consequences when using base modifications and offer design principles for the development of functional, structured RNAs in synthetic biology and therapeutic applications.
History
DepositionApr 4, 2025-
Header (metadata) releaseJul 16, 2025-
Map releaseJul 16, 2025-
UpdateJul 16, 2025-
Current statusJul 16, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_70040.png

Downloads & links

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Map

FileDownload / File: emd_70040.map.gz / Format: CCP4 / Size: 22.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
3.68 Å/pix.
x 180 pix.
= 662.4 Å		3.68 Å/pix.
x 180 pix.
= 662.4 Å		3.68 Å/pix.
x 180 pix.
= 662.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 3.68 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.0
Minimum - Maximum-0.9649189 - 6.324188
Average (Standard dev.)-0.0033620184 (±0.1927197)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions180180180
Spacing180180180
CellA=B=C: 662.4 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_70040_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_70040_half_map_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_70040_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Dimeric structure of a 6-helix bundle with a clasp helix transcri...

EntireName: Dimeric structure of a 6-helix bundle with a clasp helix transcribed with N1-methyl-pseudouridine.
Components
  • Complex: Dimeric structure of a 6-helix bundle with a clasp helix transcribed with N1-methyl-pseudouridine.
    • RNA: Twist corrected 6HBC type 2 dimer transcribed with N1-methyl-pseudouridine

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Supramolecule #1: Dimeric structure of a 6-helix bundle with a clasp helix transcri...

SupramoleculeName: Dimeric structure of a 6-helix bundle with a clasp helix transcribed with N1-methyl-pseudouridine.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 464 KDa

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Macromolecule #1: Twist corrected 6HBC type 2 dimer transcribed with N1-methyl-pseu...

MacromoleculeName: Twist corrected 6HBC type 2 dimer transcribed with N1-methyl-pseudouridine
type: rna / ID: 1
Source (natural)Organism: synthetic construct (others)
SequenceString: GGGUCCGCAC UUUGCACCGA GCUCUCGGCA ACAAGCCAGC CGAGAGUUCG GUAGUGGGUA UGUUCGCAUA CCCGCUUCCA GUACGCUAAU UCGUUAGUGU ACUCCGCUGU UAAGGGUUCG CCCUUGACAG GCCCUGAUAG UCGUUCGCGG CUAUCAGCCG GAGUGUGACU ...String:
GGGUCCGCAC UUUGCACCGA GCUCUCGGCA ACAAGCCAGC CGAGAGUUCG GUAGUGGGUA UGUUCGCAUA CCCGCUUCCA GUACGCUAAU UCGUUAGUGU ACUCCGCUGU UAAGGGUUCG CCCUUGACAG GCCCUGAUAG UCGUUCGCGG CUAUCAGCCG GAGUGUGACU UUCGAGUUGC ACUCGGGCCU GCAGUAUUUC GAUACUGUGG GCCCUGUCAC AUAACAAGGU AUCAGUUGUG UGACACGGGC UGGUCGCUAA AUCGACGAUA GCGAUCAGCG GCGUGGGUCC GUAAAAGACC GAUACGGAUC UACCGGGAGU UGCGAUCAAU GAUGGAGAUC GUAGCUCGGA GUAAAGUGUG GACCCUAAGG AUAACUCAUA CCGUGAGGGU UGCGAACCAU CAACGCGACC UUCGGCCAGG GUCGUAGGAA CGGUCUACCU ACGAUCCUCC CGAACUGAUU CGAACGUCGA ACGAAUCGGU UCCCGCCAGU GUUAUGAAGA UACCACAUAG CACUGGGAUG UUGCGGGUAC AAGGCUUGAG UGUUCGCAAU AUCACGGGAG UCUUUCGAGG CUCCCGUCGG AUCUAGGUCA UUCGUGACCU AGGUGGGCAU AUCCGCCUUC GGGCGGAUGU GGGCGUGUCU GUCCUUCGGG GCAGACACCG GGUGCACGCG GUUCGCCGCG UGUACGCCAC AUAAUCCUUU CGAGGGUUAU GUGGCUAUGA GUUGUCCUUA

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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 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 15.0 kPa / Details: 15mA in gloQube
VitrificationCryogen name: ETHANE
DetailsSEC purified in 50mM HEPES pH 8.0, 50mM KCl, 5mM 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: 59647 / 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 reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 12.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Software - details: non-uniform refinement / Number images used: 8333
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationNumber classes: 3 / Avg.num./class: 6000 / Software - Name: cryoSPARC
FSC plot (resolution estimation)

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