+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-3979 | |||||||||
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Title | Post-catalytic P complex spliceosome with 3' splice site docked | |||||||||
Map data | ||||||||||
Sample |
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Function / homology | Function and homology information U2-type post-spliceosomal complex / U2-type post-mRNA release spliceosomal complex / mRNA branch site recognition / spliceosomal complex disassembly / cellular bud site selection / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / nuclear mRNA surveillance / generation of catalytic spliceosome for first transesterification step / cis assembly of pre-catalytic spliceosome ...U2-type post-spliceosomal complex / U2-type post-mRNA release spliceosomal complex / mRNA branch site recognition / spliceosomal complex disassembly / cellular bud site selection / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / nuclear mRNA surveillance / generation of catalytic spliceosome for first transesterification step / cis assembly of pre-catalytic spliceosome / splicing factor binding / U4/U6 snRNP / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / 7-methylguanosine cap hypermethylation / Prp19 complex / snRNP binding / pICln-Sm protein complex / small nuclear ribonucleoprotein complex / pre-mRNA binding / U2-type catalytic step 1 spliceosome / SMN-Sm protein complex / spliceosomal tri-snRNP complex / mRNA cis splicing, via spliceosome / U2-type spliceosomal complex / poly(U) RNA binding / commitment complex / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / U4 snRNP / U2 snRNP / U1 snRNP / U2-type prespliceosome / precatalytic spliceosome / Formation of TC-NER Pre-Incision Complex / generation of catalytic spliceosome for second transesterification step / spliceosomal complex assembly / Gap-filling DNA repair synthesis and ligation in TC-NER / DNA replication origin binding / mRNA 5'-splice site recognition / protein K63-linked ubiquitination / mRNA 3'-splice site recognition / Dual incision in TC-NER / DNA replication initiation / spliceosomal tri-snRNP complex assembly / U5 snRNA binding / U5 snRNP / U2 snRNA binding / U6 snRNA binding / spliceosomal snRNP assembly / positive regulation of cell cycle / pre-mRNA intronic binding / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / catalytic step 2 spliceosome / nuclear periphery / RNA splicing / positive regulation of RNA splicing / spliceosomal complex / RING-type E3 ubiquitin transferase / mRNA splicing, via spliceosome / ubiquitin-protein transferase activity / metallopeptidase activity / ubiquitin protein ligase activity / RNA helicase activity / RNA helicase / DNA repair / chromatin binding / GTPase activity / mRNA binding / chromatin / GTP binding / ATP hydrolysis activity / mitochondrion / DNA binding / RNA binding / zinc ion binding / ATP binding / identical protein binding / nucleus / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) / Saccharomyces cerevisiae S288c (yeast) / Baker's yeast (brewer's yeast) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.7 Å | |||||||||
Authors | Wilkinson ME / Fica SM / Galej WP / Norman CM / Newman AJ / Nagai K | |||||||||
Funding support | United Kingdom, 2 items
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Citation | Journal: Science / Year: 2017 Title: Postcatalytic spliceosome structure reveals mechanism of 3'-splice site selection. Authors: Max E Wilkinson / Sebastian M Fica / Wojciech P Galej / Christine M Norman / Andrew J Newman / Kiyoshi Nagai / Abstract: Introns are removed from eukaryotic messenger RNA precursors by the spliceosome in two transesterification reactions-branching and exon ligation. The mechanism of 3'-splice site recognition during ...Introns are removed from eukaryotic messenger RNA precursors by the spliceosome in two transesterification reactions-branching and exon ligation. The mechanism of 3'-splice site recognition during exon ligation has remained unclear. Here we present the 3.7-angstrom cryo-electron microscopy structure of the yeast P-complex spliceosome immediately after exon ligation. The 3'-splice site AG dinucleotide is recognized through non-Watson-Crick pairing with the 5' splice site and the branch-point adenosine. After the branching reaction, protein factors work together to remodel the spliceosome and stabilize a conformation competent for 3'-splice site docking, thereby promoting exon ligation. The structure accounts for the strict conservation of the GU and AG dinucleotides at the 5' and 3' ends of introns and provides insight into the catalytic mechanism of exon ligation. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_3979.map.gz | 390.9 MB | EMDB map data format | |
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Header (meta data) | emd-3979-v30.xml emd-3979.xml | 63.2 KB 63.2 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_3979_fsc.xml | 17 KB | Display | FSC data file |
Images | emd_3979.png | 71 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-3979 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-3979 | HTTPS FTP |
-Validation report
Summary document | emd_3979_validation.pdf.gz | 323.3 KB | Display | EMDB validaton report |
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Full document | emd_3979_full_validation.pdf.gz | 322.4 KB | Display | |
Data in XML | emd_3979_validation.xml.gz | 15.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-3979 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-3979 | HTTPS FTP |
-Related structure data
Related structure data | 6exnMC 3980C M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_3979.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Voxel size | X=Y=Z: 1.12 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
+Entire : P complex spliceosome with 3' exon docked
+Supramolecule #1: P complex spliceosome with 3' exon docked
+Supramolecule #2: P complex spliceosome
+Supramolecule #3: 3' exon
+Macromolecule #1: U2 snRNA
+Macromolecule #2: U5 snRNA
+Macromolecule #3: U6 snRNA
+Macromolecule #7: Ligated exons: UBC4 mRNA
+Macromolecule #9: Intron lariat: UBC4 RNA
+Macromolecule #4: Pre-mRNA-splicing factor Prp8
+Macromolecule #5: Pre-mRNA-splicing factor SNU114
+Macromolecule #6: Pre-mRNA-splicing factor CWC16
+Macromolecule #8: Pre-mRNA-splicing factor CWC22
+Macromolecule #10: Pre-mRNA-splicing factor PRP46
+Macromolecule #11: Pre-mRNA-processing protein 45
+Macromolecule #12: Pre-mRNA-splicing factor BUD31
+Macromolecule #13: Pre-mRNA-splicing factor CWC2
+Macromolecule #14: Pre-mRNA-splicing factor SLT11
+Macromolecule #15: Pre-mRNA-splicing factor CEF1
+Macromolecule #16: Pre-mRNA-splicing factor CWC15
+Macromolecule #17: Pre-mRNA-splicing factor CWC21
+Macromolecule #18: Pre-mRNA-splicing factor CLF1
+Macromolecule #19: Pre-mRNA-splicing factor SYF1
+Macromolecule #20: Pre-mRNA-splicing factor ATP-dependent RNA helicase PRP22
+Macromolecule #21: U2 small nuclear ribonucleoprotein A'
+Macromolecule #22: Unassigned structure
+Macromolecule #23: U2 small nuclear ribonucleoprotein B''
+Macromolecule #24: Pre-mRNA-splicing factor Prp18
+Macromolecule #25: Small nuclear ribonucleoprotein-associated protein B
+Macromolecule #26: Pre-mRNA-splicing factor SLU7
+Macromolecule #27: Small nuclear ribonucleoprotein Sm D3
+Macromolecule #28: Small nuclear ribonucleoprotein E
+Macromolecule #29: Small nuclear ribonucleoprotein F
+Macromolecule #30: Small nuclear ribonucleoprotein G
+Macromolecule #31: Small nuclear ribonucleoprotein Sm D1
+Macromolecule #32: Small nuclear ribonucleoprotein Sm D2
+Macromolecule #33: Pre-mRNA-processing factor Prp17
+Macromolecule #34: Pre-mRNA-splicing factor SNT309
+Macromolecule #35: Pre-mRNA-processing factor Prp19
+Macromolecule #36: Pre-mRNA-splicing factor SYF2
+Macromolecule #37: MAGNESIUM ION
+Macromolecule #38: INOSITOL HEXAKISPHOSPHATE
+Macromolecule #39: GUANOSINE-5'-TRIPHOSPHATE
+Macromolecule #40: ZINC ION
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1.4 mg/mL | ||||||||||||
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Buffer | pH: 7.9 Component:
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Grid | Model: Quantifoil R2/2 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 7.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK III Details: 3 uL sample was applied to the grid, left for 30s, then blotted for 3s and immediately plunged into liquid ethane.. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-20 / Number grids imaged: 1 / Number real images: 2295 / Average exposure time: 12.0 sec. / Average electron dose: 47.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Calibrated defocus max: 3.0 µm / Calibrated defocus min: 0.2 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal magnification: 105000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |