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- PDB-8tvz: RNA origami 3-helix tile Traptamer -

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Basic information

Entry
Database: PDB / ID: 8tvz
TitleRNA origami 3-helix tile Traptamer
ComponentsRNA (363-MER)
KeywordsRNA / Origami / aptamer / switch / sensor / toe-hold / robot / broccoli
Function / homologyRNA / RNA (> 10) / RNA (> 100)
Function and homology information
Biological speciessynthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 5.94 Å
AuthorsMcRae, E.K.S. / Vallina, N.S. / Andersen, E.S.
Funding supportEuropean Union, Denmark, Canada, 6items
OrganizationGrant numberCountry
H2020 Marie Curie Actions of the European Commission765703European Union
Independent Research Fund Denmark - Technology and Production Sciences9040-00425B Denmark
Natural Sciences and Engineering Research Council (NSERC, Canada)532417 Canada
The Carlsberg FoundationCF20-0635 Denmark
European Research Council (ERC)683305European Union
Novo Nordisk Foundation0060694 Denmark
CitationJournal: Sci Adv / Year: 2024
Title: An RNA origami robot that traps and releases a fluorescent aptamer.
Authors: Néstor Sampedro Vallina / Ewan K S McRae / Cody Geary / Ebbe S Andersen /
Abstract: RNA nanotechnology aims to use RNA as a programmable material to create self-assembling nanodevices for application in medicine and synthetic biology. The main challenge is to develop advanced RNA ...RNA nanotechnology aims to use RNA as a programmable material to create self-assembling nanodevices for application in medicine and synthetic biology. The main challenge is to develop advanced RNA robotic devices that both sense, compute, and actuate to obtain enhanced control over molecular processes. Here, we use the RNA origami method to prototype an RNA robotic device, named the "Traptamer," that mechanically traps the fluorescent aptamer, iSpinach. The Traptamer is shown to sense two RNA key strands, acts as a Boolean AND gate, and reversibly controls the fluorescence of the iSpinach aptamer. Cryo-electron microscopy of the closed Traptamer structure at 5.45-angstrom resolution reveals the mechanical mode of distortion of the iSpinach motif. Our study suggests a general approach to distorting RNA motifs and a path forward to build sophisticated RNA machines that through sensing, computing, and actuation modules can be used to precisely control RNA functionalities in cellular systems.
History
DepositionAug 18, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 3, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
C: RNA (363-MER)


Theoretical massNumber of molelcules
Total (without water)116,9781
Polymers116,9781
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 RNA (363-MER)


Mass: 116977.742 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)

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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: 14-14 Traptamer / Type: COMPLEX
Details: An RNA origami 3 helix tile with an inactive broccoli aptamer in the central helix.
Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: synthetic construct (others)
Source (recombinant)Organism: synthetic construct (others)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
140 mMHEPES1
250 mMPotassium ChlorideKCL1
35 mMMagnesium ChlorideMgCl21
SpecimenConc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 15mA / Grid material: GOLD / Grid mesh size: 300 divisions/in.
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 294 K / Details: Protochips Au-Flat 1.2/1.3 grids.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Details: Pixel size of 0.645 A/px
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV

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Processing

EM software
IDNameCategory
2EPUimage acquisition
7UCSF ChimeraXmodel fitting
13cryoSPARC3D reconstruction
14ISOLDEmodel refinement
15PHENIXmodel refinement
16Cootmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 5.94 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 220000 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER
Atomic model buildingPDB-ID: 5OB3

5ob3


Accession code: 5OB3 / Source name: PDB / Type: experimental model

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