+Open data
-Basic information
Entry | Database: PDB / ID: 5gap | ||||||
---|---|---|---|---|---|---|---|
Title | Body region of the U4/U6.U5 tri-snRNP | ||||||
Components |
| ||||||
Keywords | TRANSCRIPTION / snRNP / spliceosome / RNA-protein complex / U4/U6.U5 snRNP | ||||||
Function / homology | Function and homology information spliceosomal conformational changes to generate catalytic conformation / snoRNA splicing / snoRNA guided rRNA 2'-O-methylation / box C/D sno(s)RNA 3'-end processing / box C/D methylation guide snoRNP complex / U4/U6 snRNP / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / U4 snRNA binding / spliceosomal tri-snRNP complex / U3 snoRNA binding ...spliceosomal conformational changes to generate catalytic conformation / snoRNA splicing / snoRNA guided rRNA 2'-O-methylation / box C/D sno(s)RNA 3'-end processing / box C/D methylation guide snoRNP complex / U4/U6 snRNP / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / U4 snRNA binding / spliceosomal tri-snRNP complex / U3 snoRNA binding / U4 snRNP / precatalytic spliceosome / Major pathway of rRNA processing in the nucleolus and cytosol / generation of catalytic spliceosome for second transesterification step / spliceosomal complex assembly / 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 / maturation of SSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / maturation of SSU-rRNA / small-subunit processome / spliceosomal complex / mRNA splicing, via spliceosome / metallopeptidase activity / nucleic acid binding / RNA helicase activity / RNA helicase / mRNA binding / nucleolus / ATP hydrolysis activity / mitochondrion / RNA binding / nucleoplasm / ATP binding / identical protein binding / nucleus / cytoplasm Similarity search - Function | ||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å | ||||||
Authors | Nguyen, T.H.D. / Galej, W.P. / Oubridge, C. / Bai, X.C. / Newman, A. / Scheres, S. / Nagai, K. | ||||||
Funding support | United Kingdom, 1items
| ||||||
Citation | Journal: Nature / Year: 2016 Title: Cryo-EM structure of the yeast U4/U6.U5 tri-snRNP at 3.7 Å resolution. Authors: Thi Hoang Duong Nguyen / Wojciech P Galej / Xiao-Chen Bai / Chris Oubridge / Andrew J Newman / Sjors H W Scheres / Kiyoshi Nagai / Abstract: U4/U6.U5 tri-snRNP represents a substantial part of the spliceosome before activation. A cryo-electron microscopy structure of Saccharomyces cerevisiae U4/U6.U5 tri-snRNP at 3.7 Å resolution led ...U4/U6.U5 tri-snRNP represents a substantial part of the spliceosome before activation. A cryo-electron microscopy structure of Saccharomyces cerevisiae U4/U6.U5 tri-snRNP at 3.7 Å resolution led to an essentially complete atomic model comprising 30 proteins plus U4/U6 and U5 small nuclear RNAs (snRNAs). The structure reveals striking interweaving interactions of the protein and RNA components, including extended polypeptides penetrating into subunit interfaces. The invariant ACAGAGA sequence of U6 snRNA, which base-pairs with the 5'-splice site during catalytic activation, forms a hairpin stabilized by Dib1 and Prp8 while the adjacent nucleotides interact with the exon binding loop 1 of U5 snRNA. Snu114 harbours GTP, but its putative catalytic histidine is held away from the γ-phosphate by hydrogen bonding to a tyrosine in the amino-terminal domain of Prp8. Mutation of this histidine to alanine has no detectable effect on yeast growth. The structure provides important new insights into the spliceosome activation process leading to the formation of the catalytic centre. | ||||||
History |
|
-Structure visualization
Movie |
Movie viewer |
---|---|
Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 5gap.cif.gz | 878.7 KB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb5gap.ent.gz | 657.9 KB | Display | PDB format |
PDBx/mmJSON format | 5gap.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 5gap_validation.pdf.gz | 509.7 KB | Display | wwPDB validaton report |
---|---|---|---|---|
Full document | 5gap_full_validation.pdf.gz | 558.8 KB | Display | |
Data in XML | 5gap_validation.xml.gz | 67.3 KB | Display | |
Data in CIF | 5gap_validation.cif.gz | 109.4 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ga/5gap ftp://data.pdbj.org/pub/pdb/validation_reports/ga/5gap | HTTPS FTP |
-Related structure data
Related structure data | 8014MC 8006C 8007C 8008C 8009C 8010C 8011C 8012C 8013C 5gamC 5ganC 5gaoC M: map data used to model this data C: citing same article (ref.) |
---|---|
Similar structure data |
-Links
-Assembly
Deposited unit |
|
---|---|
1 |
|
-Components
-RNA chain , 3 types, 3 molecules VWU
#1: RNA chain | Mass: 21528.688 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Details: U4 snRNA 5' region, nucleotides 1-67. / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: GenBank: 807071957 |
---|---|
#2: RNA chain | Mass: 35883.176 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: GenBank: 807071964 |
#3: RNA chain | Mass: 68643.344 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: GenBank: 807071959 |
-Protein , 5 types, 5 molecules xDFKB
#4: Protein | Mass: 6996.616 Da / Num. of mol.: 1 / Source method: isolated from a natural source Details: Clear helical regions were built as poly(Ala) but could not be assigned to a specific protein. Source: (natural) Saccharomyces cerevisiae (brewer's yeast) |
---|---|
#8: Protein | Mass: 16798.387 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q06819 |
#9: Protein | Mass: 56382.516 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P49704 |
#11: Protein | Mass: 13582.855 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P39990 |
#12: Protein | Mass: 246470.266 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P32639, RNA helicase |
-Pre-mRNA-splicing factor ... , 2 types, 2 molecules AJ
#5: Protein | Mass: 279867.469 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P33334 |
---|---|
#7: Protein | Mass: 104370.133 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P19735 |
-U4/U6 small nuclear ribonucleoprotein ... , 2 types, 2 molecules HG
#6: Protein | Mass: 52506.984 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P20053 |
---|---|
#10: Protein | Mass: 55974.320 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q03338 |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Body region of the U4/U6.U5 tri-snRNP complex / Type: COMPLEX / Entity ID: all / Source: NATURAL |
---|---|
Molecular weight | Value: 0.5 MDa / Experimental value: NO |
Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Buffer solution | pH: 7.9 |
Buffer component | Conc.: 1 mM / Name: DTT |
Specimen | Conc.: 0.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: 3.5 microlitre of sample was applied to grid. |
Specimen support | Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K Details: Grids were blotted at 4 deg C for 2 seconds before plunging. |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
---|---|
Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 81000 X / Calibrated magnification: 35714 X / Nominal defocus max: 3500 nm / Nominal defocus min: 500 nm / Cs: 2 mm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 16 sec. / Electron dose: 38 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 2477 |
EM imaging optics | Energyfilter name: GIF Quantum |
Image scans | Movie frames/image: 20 / Used frames/image: 1-20 |
-Processing
Software | Name: REFMAC / Version: 5.8.0124 / Classification: refinement | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
EM software |
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING ONLY | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 473827 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 140155 / Num. of class averages: 1 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: AB INITIO MODEL / Space: RECIPROCAL | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement | Resolution: 3.6→180.18 Å / Cor.coef. Fo:Fc: 0.975 / SU B: 24.876 / SU ML: 0.368 / ESU R: 0.688 Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Solvent computation | Solvent model: PARAMETERS FOR MASK CACLULATION | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 316.59 Å2
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement step | Cycle: 1 / Total: 31573 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
|