+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-13630 | |||||||||
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Title | Mature conformer of a 6-helix bundle of RNA with clasp | |||||||||
Map data | ||||||||||
Sample |
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Keywords | RNA / origami / nanostructure | |||||||||
Biological species | synthetic construct (others) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.9 Å | |||||||||
Authors | McRae EKS / Bogglid A / Boesen T / Andersen ES | |||||||||
Funding support | Denmark, 1 items
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Citation | Journal: Nat Nanotechnol / Year: 2023 Title: Structure, folding and flexibility of co-transcriptional RNA origami. Authors: Ewan K S McRae / Helena Østergaard Rasmussen / Jianfang Liu / Andreas Bøggild / Michael T A Nguyen / Nestor Sampedro Vallina / Thomas Boesen / Jan Skov Pedersen / Gang Ren / Cody Geary / ...Authors: Ewan K S McRae / Helena Østergaard Rasmussen / Jianfang Liu / Andreas Bøggild / Michael T A Nguyen / Nestor Sampedro Vallina / Thomas Boesen / Jan Skov Pedersen / Gang Ren / Cody Geary / Ebbe Sloth Andersen / Abstract: RNA origami is a method for designing RNA nanostructures that can self-assemble through co-transcriptional folding with applications in nanomedicine and synthetic biology. However, to advance the ...RNA origami is a method for designing RNA nanostructures that can self-assemble through co-transcriptional folding with applications in nanomedicine and synthetic biology. However, to advance the method further, an improved understanding of RNA structural properties and folding principles is required. Here we use cryogenic electron microscopy to study RNA origami sheets and bundles at sub-nanometre resolution revealing structural parameters of kissing-loop and crossover motifs, which are used to improve designs. In RNA bundle designs, we discover a kinetic folding trap that forms during folding and is only released after 10 h. Exploration of the conformational landscape of several RNA designs reveal the flexibility of helices and structural motifs. Finally, sheets and bundles are combined to construct a multidomain satellite shape, which is characterized by individual-particle cryo-electron tomography to reveal the domain flexibility. Together, the study provides a structural basis for future improvements to the design cycle of genetically encoded RNA nanodevices. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_13630.map.gz | 54.7 MB | EMDB map data format | |
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Header (meta data) | emd-13630-v30.xml emd-13630.xml | 21.4 KB 21.4 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_13630_fsc.xml | 10.7 KB | Display | FSC data file |
Images | emd_13630.png | 46.1 KB | ||
Masks | emd_13630_msk_1.map | 111.5 MB | Mask map | |
Filedesc metadata | emd-13630.cif.gz | 6.1 KB | ||
Others | emd_13630_additional_1.map.gz emd_13630_half_map_1.map.gz emd_13630_half_map_2.map.gz | 104.7 MB 103.2 MB 103.2 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-13630 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13630 | HTTPS FTP |
-Validation report
Summary document | emd_13630_validation.pdf.gz | 732.4 KB | Display | EMDB validaton report |
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Full document | emd_13630_full_validation.pdf.gz | 731.9 KB | Display | |
Data in XML | emd_13630_validation.xml.gz | 18.7 KB | Display | |
Data in CIF | emd_13630_validation.cif.gz | 24.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13630 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13630 | HTTPS FTP |
-Related structure data
Related structure data | 7ptlMC 7ptkC 7ptqC 7ptsC 7qduC M: atomic model generated by this map C: citing same article (ref.) |
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-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_13630.map.gz / Format: CCP4 / Size: 111.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.32 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_13630_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Sharpened map from cyroSPARC with bfactor of 250.
File | emd_13630_additional_1.map | ||||||||||||
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Annotation | Sharpened map from cyroSPARC with bfactor of 250. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_13630_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_13630_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Mature conformer of a 6-helix bundle of RNA with clasp
Entire | Name: Mature conformer of a 6-helix bundle of RNA with clasp |
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Components |
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-Supramolecule #1: Mature conformer of a 6-helix bundle of RNA with clasp
Supramolecule | Name: Mature conformer of a 6-helix bundle of RNA with clasp type: complex / ID: 1 / Parent: 0 / Macromolecule list: all Details: In vitro transcribed RNA purified by SEC. This is the mature conformer. |
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Source (natural) | Organism: synthetic construct (others) |
Molecular weight | Theoretical: 232 KDa |
-Macromolecule #1: Chains: B
Macromolecule | Name: Chains: B / type: rna / ID: 1 / Number of copies: 1 |
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Source (natural) | Organism: synthetic construct (others) |
Molecular weight | Theoretical: 231.958125 KDa |
Sequence | String: GGGAGAGUAC UAUUCAGAUG CAGACCGCAA GUUCAGAGCG GUUUGCAUCU AGGGUACGUU UUCGAACGUA UCCUCCGACU AAGUGUAUU CGUAUACUUA GUGCCUUGUG CCUGCUUCGG CAGGCAUGAC CCAAAUGUGC CUUUCGGGGC ACAUUUCCGG U CAUCCAAG ...String: GGGAGAGUAC UAUUCAGAUG CAGACCGCAA GUUCAGAGCG GUUUGCAUCU AGGGUACGUU UUCGAACGUA UCCUCCGACU AAGUGUAUU CGUAUACUUA GUGCCUUGUG CCUGCUUCGG CAGGCAUGAC CCAAAUGUGC CUUUCGGGGC ACAUUUCCGG U CAUCCAAG UUCGCUUGGG UGAUGCGGGC GUAUAGGUUC GUCUAUACGU CCGCGUUUUC CGAGAAGAGG UAACUCGGGA AA CCGGUCC ACGUGACAAA GGUAGAGUUA CGUGGAGGGA GCAGCUGCAA AGGGAUAAUG CAGUUGCUGG CUGGAUGCCA GAA CUCACG ACUGGCAUCU ACGGGGAUGG UGCUCUCCCA AUUCUCCAUU UACCGCCGAA UCGACCCCAA CGUGAGAGGG GUCG GUUCC CCGAGCAUAG ACCAAUAUCC CAGGUUUAUG CUCCCCAACG CUGGACGAAC UACCUACGUC UAGCGUUCCG GCAAA UGAG UCAAUACCUC AGACUUAUUU GCGGUGCCUG AGCCUAAACU GAACAUGGGU UCAGGCAUCU UGGCUCCAGU UCGCUG GAG CCGACGGUAG CGCUGCGUUC GCGCAGUGCU AGGGAGCAUC CGUUUUCGAG CGGAUGCUGG GCGGUUGCCU GUUCGCA GG CAAUCGGGCC UACUCAUGAU UCGUCAUGAG UGGUGACAGC GUGAUGUUCG CAUUACGCUG UCGGGUAGAU GGAGAAUU |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 2.3 mg/mL | ||||||||||||
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Buffer | pH: 8 Component:
Details: Freshly prepared and filtered through 0.22 micron filter prior to use. | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 99 % / Chamber temperature: 294 K / Instrument: LEICA EM GP Details: 3 microlitre droplet, 4 second delay before blotting, 6 second blot, 0 second delay before plunging.. | ||||||||||||
Details | Sample was purified by size exclusion chromatography and concentrated in an Amicon spin concentrator. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 8359 / Average exposure time: 1.5 sec. / Average electron dose: 60.0 e/Å2 / Details: Images were collected as 56 frame movies. |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.75 µm / Nominal magnification: 130000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Source name: PDB / Chain - Initial model type: experimental model |
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Details | Helical templates and tetraloops were generated in RNAbuild. Kissing loops were modelling based on the PDB:2D1B structure. A combination of PHENIX RSR and ISOLDE were used to arrive at the final structure. |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT / Overall B value: 303 / Target criteria: Correlation Coefficient |
Output model | PDB-7ptl: |