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Open data
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Basic information
Entry | Database: PDB / ID: 9h86 | ||||||
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Title | Small circular RNA dimer - Class 4 | ||||||
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![]() | RNA / small RNA circles / dimer / RNA crossover | ||||||
Function / homology | RNA / RNA (> 10)![]() | ||||||
Biological species | synthetic construct (others) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 9.3 Å | ||||||
![]() | McRae, E.K. / Kristoffersen, E.L. / Holliger, P. / Andersen, E.S. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Roles of dimeric intermediates in RNA-catalyzed rolling circle synthesis. Authors: Emil L Kristoffersen / Ewan K S McRae / Niels R Sørensen / Philipp Holliger / Ebbe S Andersen / ![]() ![]() ![]() Abstract: The RNA world hypothesis is supported by the discovery of RNA polymerase ribozymes that can perform RNA-catalyzed RNA replication processes on different RNA templates. Recently, RNA-catalyzed rolling ...The RNA world hypothesis is supported by the discovery of RNA polymerase ribozymes that can perform RNA-catalyzed RNA replication processes on different RNA templates. Recently, RNA-catalyzed rolling circle synthesis (RCS) on small circular RNA (scRNA) templates has been demonstrated. However, the structural and dynamic properties of scRNA replication and its products and intermediates have not been explored. Here, we have used cryogenic electron microscopy (cryo-EM) to characterize products and intermediates relevant for RCS replication. We find that these form an unexpectedly diverse group of RNA nanostructures. The main structural motif observed is a fully hybridized dimeric complex composed of two scRNAs and their complement strands resolved to 5.3 Å. Cryo-EM also reveals higher-order dimer filaments and dimer assembly intermediates, suggesting an assembly mechanism for the observed complexes. We show that the dimer complexes are stable and inhibit RNA-catalyzed RCS but can be reactivated by addition of more scRNA templates. We propose dimer formation as a general property of RCS replication and speculate that dimers might have benefited a primordial RNA genetic system by providing a stable ''storage'' form for RNA replication products and by coordinated RNA replication on both scRNA template strands. | ||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 210.1 KB | Display | ![]() |
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PDB format | ![]() | 156.1 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 985.4 KB | Display | ![]() |
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Full document | ![]() | 990.9 KB | Display | |
Data in XML | ![]() | 23.1 KB | Display | |
Data in CIF | ![]() | 33 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 51932MC ![]() 8s6wC ![]() 9h82C ![]() 9h83C ![]() 9h8aC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: RNA chain | Mass: 11817.235 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) synthetic construct (others) / Production host: synthetic construct (others) #2: RNA chain | Mass: 11191.466 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Details: Circular RNA / Source: (gene. exp.) synthetic construct (others) / Production host: synthetic construct (others) Has protein modification | N | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Dimer intermediate class 2 composed of 2 circular RNA and 2 complimentary RNA Type: COMPLEX / Details: Dimer prepared by annealing / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: synthetic construct (others) |
Source (recombinant) | Organism: synthetic construct (others) |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 90 % / Chamber temperature: 283 K |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: TFS KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 1800 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 1.5 sec. / Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 2 / Num. of real images: 6820 |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 668921 | ||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 9.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 26859 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 231 / Protocol: OTHER / Space: REAL | ||||||||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE |