+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-13636 | |||||||||
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Title | RNA origami 5-helix tile | |||||||||
Map data | ||||||||||
Sample |
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Keywords | RNA / origami / nanostructure | |||||||||
Biological species | synthetic construct (others) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 5.71 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_13636.map.gz | 4.1 MB | EMDB map data format | |
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Header (meta data) | emd-13636-v30.xml emd-13636.xml | 21.1 KB 21.1 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_13636_fsc.xml | 4.4 KB | Display | FSC data file |
Images | emd_13636.png | 46.8 KB | ||
Masks | emd_13636_msk_1.map | 8 MB | Mask map | |
Filedesc metadata | emd-13636.cif.gz | 5.9 KB | ||
Others | emd_13636_additional_1.map.gz emd_13636_half_map_1.map.gz emd_13636_half_map_2.map.gz | 7.6 MB 7.4 MB 7.4 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-13636 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13636 | HTTPS FTP |
-Validation report
Summary document | emd_13636_validation.pdf.gz | 685.8 KB | Display | EMDB validaton report |
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Full document | emd_13636_full_validation.pdf.gz | 685.4 KB | Display | |
Data in XML | emd_13636_validation.xml.gz | 10.9 KB | Display | |
Data in CIF | emd_13636_validation.cif.gz | 13.6 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13636 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13636 | HTTPS FTP |
-Related structure data
Related structure data | 7ptsMC 7ptkC 7ptlC 7ptqC 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_13636.map.gz / Format: CCP4 / Size: 8 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: 2.59 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_13636_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Sharpened map from cryoSPARC with bfactor 306.
File | emd_13636_additional_1.map | ||||||||||||
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Annotation | Sharpened map from cryoSPARC with bfactor 306. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_13636_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_13636_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : 5 helix tile B
Entire | Name: 5 helix tile B |
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Components |
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-Supramolecule #1: 5 helix tile B
Supramolecule | Name: 5 helix tile B / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all / Details: In vitro transcribed RNA purified by SEC. |
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Source (natural) | Organism: synthetic construct (others) |
Molecular weight | Theoretical: 180 KDa |
-Macromolecule #1: 5HT-B
Macromolecule | Name: 5HT-B / type: rna / ID: 1 / Details: In vitro transcribed RNA / Number of copies: 1 |
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Source (natural) | Organism: synthetic construct (others) |
Molecular weight | Theoretical: 179.904219 KDa |
Sequence | String: GGAACUUAUU CAUUAUACCC CUCAUUUUCG AGUGAGGCGG AUUAGAUUCG UCUGAUCGUA GUGGUGGUGG CUUGCUUCGG CGAGCCCGG CAGGCCUUCG GGCUUGCUCA GGGGUUCGCU CCUGAGAUUA UCAUCCAACC UCCAAGGAUG GUAAUCCGGC G UCGAUUGU ...String: GGAACUUAUU CAUUAUACCC CUCAUUUUCG AGUGAGGCGG AUUAGAUUCG UCUGAUCGUA GUGGUGGUGG CUUGCUUCGG CGAGCCCGG CAGGCCUUCG GGCUUGCUCA GGGGUUCGCU CCUGAGAUUA UCAUCCAACC UCCAAGGAUG GUAAUCCGGC G UCGAUUGU AAGGUCUAAA CAAUUGACGC ACCACUACUA CAGUCCCAAC GACUGAGGGA CUCGUCGACU CGGUUAAGAU CG GAAACCG GGUCGAGGUA UAGUGAAUGA GUUCCAAUUC UCCAUUUACG GCAUCUUGUG CGAACCGAUC ACGCACGAGA UGG UCCUUA CGGGGUGUCC CAACAGUCGA GGGGCACGCA GGAUGCCUAG AAUAGACCAC UAGGUAUCCU CAGUGCGCCA UGCA AUGGA GGAGCAUGGU GCACUGCAGG GCUCCUGAAA AGGAGUCCUG GCAGAAGGGU UCGCCUUUCU CCCGUAGGGA CGACU UUCU UUGAAAAAAG GAAGUCCCAG UUUGCUUCGG CGAACUCCGA CUCUAGGAAA CUAGGGUCGG GUAGAUGGAG AAUU |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 3.5 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: 1331 / 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: 306 / Target criteria: Correlation Coefficient |
Output model | PDB-7pts: |