+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-4658 | |||||||||
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Title | Structure of the human U5.U4/U6 tri-snRNP at 2.9A resolution. | |||||||||
Map data | Human U4.U5/U6 tri-snRNP, overall map. | |||||||||
Sample |
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Function / homology | Function and homology information Lsm2-8 complex / U6 snRNA 3'-end binding / spliceosomal snRNP complex / ribonucleoprotein complex localization / U4atac snRNP / positive regulation of cytotoxic T cell differentiation / maturation of 5S rRNA / RNA localization / R-loop processing / U4atac snRNA binding ...Lsm2-8 complex / U6 snRNA 3'-end binding / spliceosomal snRNP complex / ribonucleoprotein complex localization / U4atac snRNP / positive regulation of cytotoxic T cell differentiation / maturation of 5S rRNA / RNA localization / R-loop processing / U4atac snRNA binding / mRNA decay by 5' to 3' exoribonuclease / Lsm1-7-Pat1 complex / U6 snRNP / box C/D sno(s)RNA binding / PH domain binding / dense fibrillar component / U2 snRNP binding / U7 snRNA binding / histone pre-mRNA DCP binding / U7 snRNP / histone pre-mRNA 3'end processing complex / cis assembly of pre-catalytic spliceosome / SLBP independent Processing of Histone Pre-mRNAs / SLBP Dependent Processing of Replication-Dependent Histone Pre-mRNAs / U4/U6 snRNP / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / protein methylation / U12-type spliceosomal complex / methylosome / 7-methylguanosine cap hypermethylation / snRNP binding / U1 snRNP binding / P-body assembly / pICln-Sm protein complex / RNA splicing, via transesterification reactions / small nuclear ribonucleoprotein complex / U2-type catalytic step 1 spliceosome / U4 snRNA binding / sno(s)RNA-containing ribonucleoprotein complex / box C/D methylation guide snoRNP complex / SMN-Sm protein complex / spliceosomal tri-snRNP complex / P granule / telomerase holoenzyme complex / mRNA cis splicing, via spliceosome / telomerase RNA binding / U2-type precatalytic spliceosome / U2-type spliceosomal complex / commitment complex / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / box C/D snoRNP assembly / U4 snRNP / rRNA modification in the nucleus and cytosol / U2 snRNP / RNA Polymerase II Transcription Termination / U3 snoRNA binding / U1 snRNP / tRNA processing / Cajal body / U2-type prespliceosome / K63-linked polyubiquitin modification-dependent protein binding / precatalytic spliceosome / spliceosomal complex assembly / mRNA Splicing - Minor Pathway / negative regulation of mRNA splicing, via spliceosome / mRNA catabolic process / nuclear-transcribed mRNA catabolic process / MLL1 complex / spliceosomal tri-snRNP complex assembly / single fertilization / Major pathway of rRNA processing in the nucleolus and cytosol / U5 snRNA binding / U5 snRNP / U2 snRNA binding / RNA processing / U6 snRNA binding / ribonucleoprotein complex binding / spliceosomal snRNP assembly / pre-mRNA intronic binding / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / catalytic step 2 spliceosome / mRNA Splicing - Major Pathway / RNA splicing / response to cocaine / helicase activity / maturation of SSU-rRNA / small-subunit processome / response to bacterium / spliceosomal complex / P-body / small GTPase binding / mRNA splicing, via spliceosome / mRNA processing / osteoblast differentiation / cellular response to xenobiotic stimulus / cellular response to tumor necrosis factor / ATPase binding / ribosomal small subunit biogenesis Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) / Human (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.92 Å | |||||||||
Authors | Charenton C / Wilkinson ME / Nagai K | |||||||||
Funding support | United Kingdom, 2 items
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Citation | Journal: Science / Year: 2019 Title: Mechanism of 5' splice site transfer for human spliceosome activation. Authors: Clément Charenton / Max E Wilkinson / Kiyoshi Nagai / Abstract: The prespliceosome, comprising U1 and U2 small nuclear ribonucleoproteins (snRNPs) bound to the precursor messenger RNA 5' splice site (5'SS) and branch point sequence, associates with the U4/U6.U5 ...The prespliceosome, comprising U1 and U2 small nuclear ribonucleoproteins (snRNPs) bound to the precursor messenger RNA 5' splice site (5'SS) and branch point sequence, associates with the U4/U6.U5 tri-snRNP to form the fully assembled precatalytic pre-B spliceosome. Here, we report cryo-electron microscopy structures of the human pre-B complex captured before U1 snRNP dissociation at 3.3-angstrom core resolution and the human tri-snRNP at 2.9-angstrom resolution. U1 snRNP inserts the 5'SS-U1 snRNA helix between the two RecA domains of the Prp28 DEAD-box helicase. Adenosine 5'-triphosphate-dependent closure of the Prp28 RecA domains releases the 5'SS to pair with the nearby U6 ACAGAGA-box sequence presented as a mobile loop. The structures suggest that formation of the 5'SS-ACAGAGA helix triggers remodeling of an intricate protein-RNA network to induce Brr2 helicase relocation to its loading sequence in U4 snRNA, enabling Brr2 to unwind the U4/U6 snRNA duplex to allow U6 snRNA to form the catalytic center of the 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_4658.map.gz | 260.2 MB | EMDB map data format | |
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Header (meta data) | emd-4658-v30.xml emd-4658.xml | 58.3 KB 58.3 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_4658_fsc.xml | 14.8 KB | Display | FSC data file |
Images | emd_4658.png | 181.4 KB | ||
Others | emd_4658_additional.map.gz | 260.1 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-4658 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-4658 | HTTPS FTP |
-Validation report
Summary document | emd_4658_validation.pdf.gz | 296.3 KB | Display | EMDB validaton report |
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Full document | emd_4658_full_validation.pdf.gz | 295.5 KB | Display | |
Data in XML | emd_4658_validation.xml.gz | 13.9 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-4658 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-4658 | HTTPS FTP |
-Related structure data
Related structure data | 6qw6MC 4665C 4672C 4673C 4674C 4675C 4676C 4686C 4687C 4688C 4689C 4690C 6qx9C M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | |
EM raw data | EMPIAR-10307 (Title: Human pre-B spliceosome and U4/U6.U5 tri-snRNP / Data size: 2.3 TB Data #1: Dataset 1 of human pre-B spliceosome; motion-corrected micrographs [micrographs - single frame] Data #2: Dataset 2 of human pre-B spliceosome; motion-corrected micrographs [micrographs - single frame] Data #3: Dataset 3 of human pre-B spliceosome; motion-corrected micrographs [micrographs - single frame] Data #4: Dataset 4 of human pre-B spliceosome; motion-corrected micrographs [micrographs - single frame] Data #5: Dataset 5 of human pre-B spliceosome; motion-corrected micrographs [micrographs - single frame] Data #6: Selected U4/U6.U5 tri-snRNP particles after Bayesian polishing [picked particles - single frame - processed] Data #7: Crude shifted preB particles [picked particles - single frame - unprocessed]) |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_4658.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Human U4.U5/U6 tri-snRNP, overall map. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.022 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Additional map: Composite map from the multi body refinement. For...
File | emd_4658_additional.map | ||||||||||||
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Annotation | Composite map from the multi body refinement. For each map voxel, the map value was taken from the body with the highest local resolution as calculated with RELION. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : Human U4/U6.U5 tri-snRNP
+Supramolecule #1: Human U4/U6.U5 tri-snRNP
+Supramolecule #2: U5.U4/U6 tri-snRNP
+Supramolecule #3: Probable ATP-dependent RNA helicase DDX23
+Macromolecule #1: U4/U6.U5 small nuclear ribonucleoprotein 27 kDa protein
+Macromolecule #3: Small nuclear ribonucleoprotein Sm D1
+Macromolecule #4: Small nuclear ribonucleoprotein Sm D2
+Macromolecule #5: Small nuclear ribonucleoprotein Sm D3
+Macromolecule #6: U4/U6 small nuclear ribonucleoprotein Prp3
+Macromolecule #7: U4/U6 small nuclear ribonucleoprotein Prp4
+Macromolecule #8: U4/U6 small nuclear ribonucleoprotein Prp31
+Macromolecule #9: NHP2-like protein 1
+Macromolecule #10: Small nuclear ribonucleoprotein-associated proteins B and B'
+Macromolecule #11: Small nuclear ribonucleoprotein E
+Macromolecule #12: Small nuclear ribonucleoprotein F
+Macromolecule #13: Small nuclear ribonucleoprotein G
+Macromolecule #15: Pre-mRNA-processing-splicing factor 8
+Macromolecule #16: U5 small nuclear ribonucleoprotein 200 kDa helicase
+Macromolecule #17: 116 kDa U5 small nuclear ribonucleoprotein component
+Macromolecule #18: Thioredoxin-like protein 4A
+Macromolecule #19: Pre-mRNA-processing factor 6
+Macromolecule #20: U5 small nuclear ribonucleoprotein 40 kDa protein
+Macromolecule #21: Probable ATP-dependent RNA helicase DDX23
+Macromolecule #23: U6 snRNA-associated Sm-like protein LSm2
+Macromolecule #24: U6 snRNA-associated Sm-like protein LSm3
+Macromolecule #25: U6 snRNA-associated Sm-like protein LSm4
+Macromolecule #26: U6 snRNA-associated Sm-like protein LSm5
+Macromolecule #27: U6 snRNA-associated Sm-like protein LSm6
+Macromolecule #28: U6 snRNA-associated Sm-like protein LSm7
+Macromolecule #29: U6 snRNA-associated Sm-like protein LSm8
+Macromolecule #30: RNA-binding protein 42
+Macromolecule #31: U4/U6.U5 tri-snRNP-associated protein 1
+Macromolecule #32: U4/U6.U5 tri-snRNP-associated protein 2
+Macromolecule #2: U4 snRNA
+Macromolecule #14: U5 snRNA
+Macromolecule #22: U6 snRNA
+Macromolecule #33: INOSITOL HEXAKISPHOSPHATE
+Macromolecule #34: MAGNESIUM ION
+Macromolecule #35: GUANOSINE-5'-TRIPHOSPHATE
+Macromolecule #36: ZINC ION
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1.3 mg/mL | ||||||||
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Buffer | pH: 7.9 Component:
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Grid | Model: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 6.0 nm / Pretreatment - Type: GLOW DISCHARGE | ||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK III / Details: Wait 30s, blot for 2s to 3s.. |
-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 K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-40 / Average exposure time: 6.0 sec. / Average electron dose: 50.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / 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 |