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Yorodumi- EMDB-12106: Yeast C complex spliceosome at 2.8 Angstrom resolution with Prp18... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-12106 | |||||||||
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Title | Yeast C complex spliceosome at 2.8 Angstrom resolution with Prp18/Slu7 bound, composite map | |||||||||
Map data | ||||||||||
Sample |
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Keywords | splicing / spliceosome / RNA / ribozyme | |||||||||
Function / homology | Function and homology information second spliceosomal transesterification activity / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / splicing factor binding / U4/U6 snRNP / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / 7-methylguanosine cap hypermethylation / Prp19 complex / pICln-Sm protein complex / snRNP binding ...second spliceosomal transesterification activity / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / splicing factor binding / U4/U6 snRNP / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / 7-methylguanosine cap hypermethylation / Prp19 complex / pICln-Sm protein complex / snRNP binding / U2-type catalytic step 1 spliceosome / small nuclear ribonucleoprotein complex / pre-mRNA binding / SMN-Sm protein complex / spliceosomal tri-snRNP 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 / generation of catalytic spliceosome for second transesterification step / spliceosomal complex assembly / regulation of RNA splicing / mRNA 5'-splice site recognition / mRNA 3'-splice site recognition / spliceosomal tri-snRNP complex assembly / U5 snRNA binding / U5 snRNP / U2 snRNA binding / U6 snRNA binding / spliceosomal snRNP assembly / pre-mRNA intronic binding / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / catalytic step 2 spliceosome / RNA splicing / helicase activity / spliceosomal complex / mRNA splicing, via spliceosome / mRNA processing / metallopeptidase activity / nucleic acid binding / RNA helicase activity / RNA helicase / GTPase activity / mRNA binding / GTP binding / ATP hydrolysis activity / RNA binding / zinc ion binding / ATP binding / identical protein binding / nucleus / metal ion binding / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.8 Å | |||||||||
Authors | Wilkinson ME / Fica SM | |||||||||
Funding support | United Kingdom, 2 items
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Citation | Journal: Mol Cell / Year: 2021 Title: Structural basis for conformational equilibrium of the catalytic spliceosome. Authors: Max E Wilkinson / Sebastian M Fica / Wojciech P Galej / Kiyoshi Nagai / Abstract: The ATPase Prp16 governs equilibrium between the branching (B/C) and exon ligation (C/P) conformations of the spliceosome. Here, we present the electron cryomicroscopy reconstruction of the ...The ATPase Prp16 governs equilibrium between the branching (B/C) and exon ligation (C/P) conformations of the spliceosome. Here, we present the electron cryomicroscopy reconstruction of the Saccharomyces cerevisiae C-complex spliceosome at 2.8 Å resolution and identify a novel C-complex intermediate (C) that elucidates the molecular basis for this equilibrium. The exon-ligation factors Prp18 and Slu7 bind to C before ATP hydrolysis by Prp16 can destabilize the branching conformation. Biochemical assays suggest that these pre-bound factors prime the C complex for conversion to C by Prp16. A complete model of the Prp19 complex (NTC) reveals how the branching factors Yju2 and Isy1 are recruited by the NTC before branching. Prp16 remodels Yju2 binding after branching, allowing Yju2 to remain tethered to the NTC in the C complex to promote exon ligation. Our results explain how Prp16 action modulates the dynamic binding of step-specific factors to alternatively stabilize the C or C conformation and establish equilibrium of the catalytic spliceosome. | |||||||||
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_12106.map.gz | 128 MB | EMDB map data format | |
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Header (meta data) | emd-12106-v30.xml emd-12106.xml | 62 KB 62 KB | Display Display | EMDB header |
Images | emd_12106.png | 202.5 KB | ||
Filedesc metadata | emd-12106.cif.gz | 17.9 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-12106 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-12106 | HTTPS FTP |
-Validation report
Summary document | emd_12106_validation.pdf.gz | 440.8 KB | Display | EMDB validaton report |
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Full document | emd_12106_full_validation.pdf.gz | 440.4 KB | Display | |
Data in XML | emd_12106_validation.xml.gz | 7.5 KB | Display | |
Data in CIF | emd_12106_validation.cif.gz | 8.6 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-12106 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-12106 | HTTPS FTP |
-Related structure data
Related structure data | 7b9vMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | |
EM raw data | EMPIAR-10687 (Title: Yeast C, Ci, C*, and P complex spliceosomes / Data size: 8.9 TB Data #1: Unaligned movies of C-complex spliceosome with 3' splice site AG to AC mutation (Dataset 1) [micrographs - multiframe] Data #2: Unaligned movies of C and C*-complex spliceosomes with 3' splice site AG to AdG mutation (Dataset 2) [micrographs - multiframe] Data #3: Unaligned movies of C and C*-complex spliceosomes with 3' splice site AG to AdG mutation (Dataset 3) [micrographs - multiframe] Data #4: Aligned movies of C-complex spliceosomes with cold-sensitive prp16-302 mutation, purified with Cwc25 (Dataset 4) [micrographs - multiframe] Data #5: Unaligned movies of C-complex spliceosomes with cold-sensitive prp16-302 mutation, purified with Cwc25 and incubated with ATP and Mg (Dataset 5) [micrographs - multiframe] Data #6: Unaligned movies of C, C*, and P-complex spliceosomes with dominant-negative Prp22 mutation K512A, purified with Slu7 (Dataset 6) [micrographs - multiframe] Data #7: Unaligned movies of P-complex spliceosomes with dominant-negative Prp22 mutation K512A, treated with anti-3'exon RNaseH oligo, purified in presence of Mg (Dataset 9) [micrographs - single frame] Data #8: Selected C-complex particles after polishing [picked particles - single frame - processed] Data #9: Selected P-complex particles after polishing [picked particles - single frame - processed] Data #10: Various signal subtractions for C- and P-complex spliceosomes [picked particles - single frame - processed]) |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_12106.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Voxel size | X=Y=Z: 1.145 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 : Yeast C complex spliceosome with Prp18/Slu7 bound
+Supramolecule #1: Yeast C complex spliceosome with Prp18/Slu7 bound
+Macromolecule #1: U2 snRNA
+Macromolecule #2: U5 snRNA
+Macromolecule #3: U6 snRNA
+Macromolecule #8: 5' exon of UBC4 mRNA
+Macromolecule #12: Branched intron and 3' exon of UBC4 pre-mRNA
+Macromolecule #4: Pre-mRNA-splicing factor 8
+Macromolecule #5: Pre-mRNA-splicing helicase BRR2
+Macromolecule #6: Pre-mRNA-splicing factor SNU114
+Macromolecule #7: Splicing factor YJU2
+Macromolecule #9: Pre-mRNA-splicing factor CWC25
+Macromolecule #10: Pre-mRNA-splicing factor ISY1
+Macromolecule #11: CWC22 isoform 1
+Macromolecule #13: BJ4_G0054360.mRNA.1.CDS.1
+Macromolecule #14: Pre-mRNA-processing protein 45
+Macromolecule #15: BUD31 isoform 1
+Macromolecule #16: Pre-mRNA-splicing factor CWC2
+Macromolecule #17: Pre-mRNA-splicing factor SLT11
+Macromolecule #18: Y55_G0042700.mRNA.1.CDS.1
+Macromolecule #19: Pre-mRNA-splicing factor CWC15
+Macromolecule #20: Pre-mRNA-splicing factor ATP-dependent RNA helicase PRP16
+Macromolecule #21: Pre-mRNA-splicing factor CWC21
+Macromolecule #22: CLF1 isoform 1
+Macromolecule #23: SYF1 isoform 1
+Macromolecule #24: HLJ1_G0053790.mRNA.1.CDS.1
+Macromolecule #25: Unassigned structure
+Macromolecule #26: BJ4_G0027490.mRNA.1.CDS.1
+Macromolecule #27: NTC20 isoform 1
+Macromolecule #28: Pre-mRNA-splicing factor 18
+Macromolecule #29: Small nuclear ribonucleoprotein-associated protein B
+Macromolecule #30: Pre-mRNA-splicing factor SLU7
+Macromolecule #31: Small nuclear ribonucleoprotein Sm D3
+Macromolecule #32: Small nuclear ribonucleoprotein E
+Macromolecule #33: Small nuclear ribonucleoprotein F
+Macromolecule #34: Small nuclear ribonucleoprotein G
+Macromolecule #35: Small nuclear ribonucleoprotein Sm D1
+Macromolecule #36: Small nuclear ribonucleoprotein Sm D2
+Macromolecule #37: CDC40 isoform 1
+Macromolecule #38: SNT309 isoform 1
+Macromolecule #39: Pre-mRNA-processing factor 19
+Macromolecule #40: Pre-mRNA-splicing factor SYF2
+Macromolecule #41: MAGNESIUM ION
+Macromolecule #42: POTASSIUM ION
+Macromolecule #43: D-chiro inositol hexakisphosphate
+Macromolecule #44: GUANOSINE-5'-TRIPHOSPHATE
+Macromolecule #45: ZINC ION
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.9 |
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Grid | Model: Quantifoil R2/2 / Support film - Material: CARBON / Support film - topology: CONTINUOUS |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number grids imaged: 9 / Number real images: 24115 / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: EMDB MAP |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 403474 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |