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- PDB-9zca: 5-methyl-cytidine twist corrected RNA origami 6-helix bundle type... -

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Basic information

Entry
Database: PDB / ID: 9zca
Title5-methyl-cytidine twist corrected RNA origami 6-helix bundle type-2 dimer
Components5-methyl-cytidine twist corrected RNA origami 6-helix bundle
KeywordsRNA / 5-methyl-cytidine / dimer
Function / homologyRNA / RNA (> 10) / RNA (> 100)
Function and homology information
Biological speciessynthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 11.5 Å
AuthorsMcRae, E.K.S. / Yadav, D.K.
Funding support United States, 1items
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 22, 2025Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 10, 2025Provider: repository / Type: Initial release
Revision 1.0Dec 10, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Dec 10, 2025Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
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Revision 1.0Dec 10, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
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Revision 1.1Jun 17, 2026Group: Data collection / Database references / Category: citation / citation_author / em_admin
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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: 5-methyl-cytidine twist corrected RNA origami 6-helix bundle
B: 5-methyl-cytidine twist corrected RNA origami 6-helix bundle


Theoretical massNumber of molelcules
Total (without water)475,7022
Polymers475,7022
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: RNA chain 5-methyl-cytidine twist corrected RNA origami 6-helix bundle


Mass: 237851.219 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
Has ligand of interestN
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: Dimeric twist-corrected 6-helix bundle with a clasp helix transcribed with 5-methyl-cytidine.
Type: COMPLEX / Entity ID: all / Source: NATURAL
Molecular weightValue: 0.464 MDa / Experimental value: NO
Source (natural)Organism: synthetic construct (others)
Buffer solutionpH: 8
Details: SEC purified in 50mM HEPES pH 8.0, 50mM KCl, 5mM MgCl2
SpecimenConc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: SEC purified in 50mM HEPES pH 8.0, 50mM KCl, 5mM MgCl2
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

MicroscopyModel: TFS GLACIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 700 nm / Cs: 2.7 mm
Image recordingElectron dose: 40 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)
EM imaging opticsEnergyfilter name: TFS Selectris X / Energyfilter slit width: 10 eV

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Processing

EM software
IDNameVersionCategory
1cryoSPARCparticle selection
2PHENIX1.21.2_5419model refinement
13cryoSPARC3D reconstruction
CTF correctionDetails: Patch CTF / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 73602 / Details: Blob picking on denoised micrographs
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 11.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 11203 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL / Target criteria: geometry minimization
Atomic model buildingPDB-ID: 9ZC9

9zc9


Accession code: 9ZC9 / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE

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