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- PDB-6qx9: Structure of a human fully-assembled precatalytic spliceosome (pr... -
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Open data
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Basic information
Entry | Database: PDB / ID: 6qx9 | |||||||||
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Title | Structure of a human fully-assembled precatalytic spliceosome (pre-B complex). | |||||||||
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![]() | SPLICING / RNP complex / RNA / protein / spliceosome | |||||||||
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 / Cajal body / 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 / pre-mRNA intronic binding Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.28 Å | |||||||||
![]() | Charenton, C. / Wilkinson, M.E. / 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 | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 3.2 MB | Display | ![]() |
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PDB format | ![]() | Display | ![]() | |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 4665MC ![]() 4658C ![]() 4672C ![]() 4673C ![]() 4674C ![]() 4675C ![]() 4676C ![]() 4686C ![]() 4687C ![]() 4688C ![]() 4689C ![]() 4690C ![]() 6qw6C M: map data used to model this data C: citing same article ( |
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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
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Assembly
Deposited unit | ![]()
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Components
+RNA chain , 6 types, 6 molecules 16I524
+U5 small nuclear ribonucleoprotein ... , 2 types, 2 molecules 5O5B
+Splicing factor 3B subunit ... , 5 types, 5 molecules B4B2B5B3B1
+Small nuclear ribonucleoprotein ... , 6 types, 24 molecules 135323435e4e1e2e411151211f2f5f4f224252121g2g5g4g
+U4/U6 small nuclear ribonucleoprotein ... , 3 types, 3 molecules 4B4C4A
+U1 small nuclear ribonucleoprotein ... , 3 types, 3 molecules 1K1A1C
+Protein , 11 types, 14 molecules RX5C5X1b2b5b4b5J4D5A5DBPK
+U6 snRNA-associated Sm-like protein ... , 7 types, 7 molecules 66676263686465
+U2 small nuclear ribonucleoprotein ... , 2 types, 2 molecules 2B2A
+Splicing factor 3A subunit ... , 3 types, 3 molecules A2A3A1
+U4/U6.U5 tri-snRNP-associated protein ... , 2 types, 2 molecules SU
+Non-polymers , 4 types, 9 molecules ![](data/chem/img/ZN.gif)
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
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Molecular weight | Value: 3.0 MDa / Experimental value: NO | ||||||||||||||||||||||||||||||
Source (natural) |
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Source (recombinant) |
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Buffer solution | pH: 7.9 | ||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 1.4 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R2/2 | ||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 6 sec. / Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
Image scans | Movie frames/image: 40 / Used frames/image: 1-40 |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1130953 | ||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.28 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 86146 / Algorithm: FOURIER SPACE / Symmetry type: POINT |