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- EMDB-4665: Structure of a human fully-assembled precatalytic spliceosome (pr... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-4665 | |||||||||
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Title | Structure of a human fully-assembled precatalytic spliceosome (pre-B complex). | |||||||||
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Function / homology | ![]() mRNA Splicing - Minor Pathway / negative regulation of protein refolding / mRNA Splicing - Major Pathway / 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 ...mRNA Splicing - Minor Pathway / negative regulation of protein refolding / mRNA Splicing - Major Pathway / 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 / positive regulation of hippo signaling / RNA localization / R-loop processing / U4atac snRNA binding / regulation of ATP-dependent activity / mRNA decay by 5' to 3' exoribonuclease / Lsm1-7-Pat1 complex / regulation of mitotic cell cycle spindle assembly checkpoint / U6 snRNP / box C/D sno(s)RNA binding / U11/U12 snRNP / PH domain binding / dense fibrillar component / U2 snRNP binding / U7 snRNA binding / histone pre-mRNA DCP binding / U7 snRNP / B-WICH complex / 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 / splicing factor binding / 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 / negative regulation of chaperone-mediated autophagy / 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 / positive regulation of mRNA splicing, via spliceosome / box C/D snoRNP assembly / U4 snRNP / rRNA modification in the nucleus and cytosol / U2 snRNP / RNA Polymerase II Transcription Termination / SAGA complex / U3 snoRNA binding / positive regulation of transcription by RNA polymerase III / U1 snRNP / pre-mRNA 5'-splice site binding / tRNA processing / U2-type prespliceosome / nuclear androgen receptor binding / K63-linked polyubiquitin modification-dependent protein binding / precatalytic spliceosome / positive regulation of transcription by RNA polymerase I / spliceosomal complex assembly / mRNA Splicing - Minor Pathway / regulation of RNA splicing / negative regulation of mRNA splicing, via spliceosome / mRNA catabolic process / mRNA 5'-splice site recognition / nuclear-transcribed mRNA catabolic process / mRNA 3'-splice site recognition / 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 / regulation of DNA repair / Cajal body / pre-mRNA intronic binding Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.28 Å | |||||||||
![]() | Charenton C / Wilkinson ME / Nagai K | |||||||||
Funding support | ![]()
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![]() | ![]() 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 |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 260.2 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 80.9 KB 80.9 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 14.9 KB | Display | ![]() |
Images | ![]() | 125.9 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6qx9MC ![]() 4658C ![]() 4672C ![]() 4673C ![]() 4674C ![]() 4675C ![]() 4676C ![]() 4686C ![]() 4687C ![]() 4688C ![]() 4689C ![]() 4690C ![]() 6qw6C C: citing same article ( M: atomic model generated by this map |
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Similar structure data | |
EM raw data | ![]() 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]) |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | None | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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
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Sample components
+Entire : Fully-assembled precatalytic spliceosome (pre-B complex).
+Supramolecule #1: Fully-assembled precatalytic spliceosome (pre-B complex).
+Supramolecule #2: Fully-assembled precatalytic spliceosome (pre-B complex).
+Supramolecule #3: Probable ATP-dependent RNA helicase DDX23
+Supramolecule #4: AdML pre-mRNA
+Macromolecule #1: U1 snRNA
+Macromolecule #2: U6 snRNA
+Macromolecule #8: AdML pre-mRNA
+Macromolecule #17: U5 snRNA
+Macromolecule #32: U2 snRNA
+Macromolecule #45: U4 snRNA
+Macromolecule #3: U5 small nuclear ribonucleoprotein 40 kDa protein
+Macromolecule #4: Splicing factor 3B subunit 4
+Macromolecule #5: Small nuclear ribonucleoprotein Sm D3
+Macromolecule #6: U4/U6 small nuclear ribonucleoprotein Prp4
+Macromolecule #7: Small nuclear ribonucleoprotein E
+Macromolecule #9: U1 small nuclear ribonucleoprotein 70 kDa
+Macromolecule #10: U4/U6 small nuclear ribonucleoprotein Prp31
+Macromolecule #11: Small nuclear ribonucleoprotein Sm D1
+Macromolecule #12: RNA-binding protein 42
+Macromolecule #13: Small nuclear ribonucleoprotein F
+Macromolecule #14: U6 snRNA-associated Sm-like protein LSm6
+Macromolecule #15: U4/U6.U5 small nuclear ribonucleoprotein 27 kDa protein
+Macromolecule #16: Small nuclear ribonucleoprotein Sm D2
+Macromolecule #18: U6 snRNA-associated Sm-like protein LSm7
+Macromolecule #19: U6 snRNA-associated Sm-like protein LSm2
+Macromolecule #20: U2 small nuclear ribonucleoprotein B''
+Macromolecule #21: Splicing factor 3A subunit 2
+Macromolecule #22: Splicing factor 3B subunit 2
+Macromolecule #23: 116 kDa U5 small nuclear ribonucleoprotein component
+Macromolecule #24: Probable ATP-dependent RNA helicase DDX23
+Macromolecule #25: Small nuclear ribonucleoprotein-associated proteins B and B'
+Macromolecule #26: Splicing factor 3B subunit 5
+Macromolecule #27: U1 small nuclear ribonucleoprotein A
+Macromolecule #28: U4/U6.U5 tri-snRNP-associated protein 1
+Macromolecule #29: Pre-mRNA-processing factor 6
+Macromolecule #30: NHP2-like protein 1
+Macromolecule #31: U6 snRNA-associated Sm-like protein LSm3
+Macromolecule #33: Splicing factor 3B subunit 3
+Macromolecule #34: Small nuclear ribonucleoprotein G
+Macromolecule #35: U6 snRNA-associated Sm-like protein LSm8
+Macromolecule #36: Pre-mRNA-processing-splicing factor 8
+Macromolecule #37: Splicing factor 3A subunit 3
+Macromolecule #38: U4/U6.U5 tri-snRNP-associated protein 2
+Macromolecule #39: Thioredoxin-like protein 4A
+Macromolecule #40: U6 snRNA-associated Sm-like protein LSm4
+Macromolecule #41: PHD finger-like domain-containing protein 5A
+Macromolecule #42: U1 small nuclear ribonucleoprotein C
+Macromolecule #43: Serine/threonine-protein kinase PRP4 homolog
+Macromolecule #44: U4/U6 small nuclear ribonucleoprotein Prp3
+Macromolecule #46: U2 small nuclear ribonucleoprotein A'
+Macromolecule #47: Splicing factor 3A subunit 1,Splicing factor 3A subunit 1,Splicin...
+Macromolecule #48: U6 snRNA-associated Sm-like protein LSm5
+Macromolecule #49: U5 small nuclear ribonucleoprotein 200 kDa helicase
+Macromolecule #50: Splicing factor 3B subunit 1
+Macromolecule #51: ZINC ION
+Macromolecule #52: MAGNESIUM ION
+Macromolecule #53: GUANOSINE-5'-TRIPHOSPHATE
+Macromolecule #54: INOSITOL HEXAKISPHOSPHATE
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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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: 6.0 nm / Pretreatment - Type: GLOW DISCHARGE | ||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK III |
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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: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |