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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-3683 | |||||||||
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Title | Structure of a pre-catalytic spliceosome (B2 map) | |||||||||
![]() | unsharpened B2 map | |||||||||
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Function / homology | ![]() spliceosomal conformational changes to generate catalytic conformation / mRNA decay by 5' to 3' exoribonuclease / snoRNA splicing / maturation of 5S rRNA / snoRNA guided rRNA 2'-O-methylation / Lsm1-7-Pat1 complex / U6 snRNP / box C/D sno(s)RNA 3'-end processing / deadenylation-dependent decapping of nuclear-transcribed mRNA / generation of catalytic spliceosome for first transesterification step ...spliceosomal conformational changes to generate catalytic conformation / mRNA decay by 5' to 3' exoribonuclease / snoRNA splicing / maturation of 5S rRNA / snoRNA guided rRNA 2'-O-methylation / Lsm1-7-Pat1 complex / U6 snRNP / box C/D sno(s)RNA 3'-end processing / deadenylation-dependent decapping of nuclear-transcribed mRNA / generation of catalytic spliceosome for first transesterification step / splicing factor binding / U4/U6 snRNP / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / 7-methylguanosine cap hypermethylation / snRNP binding / P-body assembly / pICln-Sm protein complex / small nuclear ribonucleoprotein complex / U4 snRNA binding / sno(s)RNA-containing ribonucleoprotein complex / box C/D methylation guide snoRNP complex / SMN-Sm protein complex / spliceosomal tri-snRNP complex / mRNA cis splicing, via spliceosome / U2-type spliceosomal complex / poly(U) RNA binding / commitment complex / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / U4 snRNP / U2 snRNP / U3 snoRNA binding / U1 snRNP / tRNA processing / U2-type prespliceosome / 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 / nuclear-transcribed mRNA catabolic process / mRNA 3'-splice site recognition / spliceosomal tri-snRNP complex assembly / U5 snRNA binding / U5 snRNP / U2 snRNA binding / U6 snRNA binding / spliceosomal snRNP assembly / cellular response to glucose starvation / 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 / P-body / mRNA splicing, via spliceosome / rRNA processing / metallopeptidase activity / nucleic acid binding / RNA helicase activity / RNA helicase / response to xenobiotic stimulus / GTPase activity / mRNA binding / nucleolus / GTP binding / ATP hydrolysis activity / mitochondrion / RNA binding / zinc ion binding / nucleoplasm / ATP binding / identical protein binding / nucleus / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 7.2 Å | |||||||||
![]() | Plaschka C / Lin P-C / Nagai K | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structure of a pre-catalytic spliceosome. Authors: Clemens Plaschka / Pei-Chun Lin / Kiyoshi Nagai / ![]() Abstract: Intron removal requires assembly of the spliceosome on precursor mRNA (pre-mRNA) and extensive remodelling to form the spliceosome's catalytic centre. Here we report the cryo-electron microscopy ...Intron removal requires assembly of the spliceosome on precursor mRNA (pre-mRNA) and extensive remodelling to form the spliceosome's catalytic centre. Here we report the cryo-electron microscopy structure of the yeast Saccharomyces cerevisiae pre-catalytic B complex spliceosome at near-atomic resolution. The mobile U2 small nuclear ribonucleoprotein particle (snRNP) associates with U4/U6.U5 tri-snRNP through the U2/U6 helix II and an interface between U4/U6 di-snRNP and the U2 snRNP SF3b-containing domain, which also transiently contacts the helicase Brr2. The 3' region of the U2 snRNP is flexibly attached to the SF3b-containing domain and protrudes over the concave surface of tri-snRNP, where the U1 snRNP may reside before its release from the pre-mRNA 5' splice site. The U6 ACAGAGA sequence forms a hairpin that weakly tethers the 5' splice site. The B complex proteins Prp38, Snu23 and Spp381 bind the Prp8 N-terminal domain and stabilize U6 ACAGAGA stem-pre-mRNA and Brr2-U4 small nuclear RNA interactions. These results provide important insights into the events leading to active site formation. | |||||||||
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 | ![]() | 392 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 71.5 KB 71.5 KB | Display Display | ![]() |
Images | ![]() | 45.8 KB | ||
Others | ![]() | 394 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 5nrlMC ![]() 3682C ![]() 3684C ![]() 3685C ![]() 3686C ![]() 3687C ![]() 3688C C: citing same article ( M: atomic model generated by this map |
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Similar structure data | |
EM raw data | ![]() Data #1: Polished parcitles [picked particles - single frame - processed]) |
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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 | unsharpened B2 map | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.43 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Additional map: sharpened B2 map
File | emd_3683_additional.map | ||||||||||||
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Annotation | sharpened B2 map | ||||||||||||
Projections & Slices |
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Density Histograms |
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Sample components
+Entire : Pre-catalytic B complex Spliceosome
+Supramolecule #1: Pre-catalytic B complex Spliceosome
+Macromolecule #1: U2 snRNA
+Macromolecule #3: U4 snRNA
+Macromolecule #4: U5 snRNA
+Macromolecule #5: U6 snRNA
+Macromolecule #16: Yeast UBC4 gene for ubiquitin-conjugating enzyme
+Macromolecule #2: U6 snRNA-associated Sm-like protein LSm3
+Macromolecule #6: U6 snRNA-associated Sm-like protein LSm7
+Macromolecule #7: U6 snRNA-associated Sm-like protein LSm8
+Macromolecule #8: Pre-mRNA-splicing factor 8
+Macromolecule #9: Pre-mRNA-splicing helicase BRR2
+Macromolecule #10: Pre-mRNA-splicing factor SNU114
+Macromolecule #11: Spliceosomal protein DIB1
+Macromolecule #12: 66 kDa U4/U6.U5 small nuclear ribonucleoprotein component
+Macromolecule #13: Pre-mRNA-processing factor 31
+Macromolecule #14: U4/U6 small nuclear ribonucleoprotein PRP3
+Macromolecule #15: U4/U6 small nuclear ribonucleoprotein PRP4
+Macromolecule #17: Pre-mRNA-splicing factor 6
+Macromolecule #18: 13 kDa ribonucleoprotein-associated protein
+Macromolecule #19: 23 kDa U4/U6.U5 small nuclear ribonucleoprotein component
+Macromolecule #20: Pre-mRNA-splicing factor 38
+Macromolecule #21: Pre-mRNA-splicing factor SPP381
+Macromolecule #22: U2 snRNP component HSH155
+Macromolecule #23: Pre-mRNA-splicing factor RSE1
+Macromolecule #24: Cold sensitive U2 snRNA suppressor 1
+Macromolecule #25: Protein HSH49
+Macromolecule #26: Pre-mRNA-splicing factor RDS3
+Macromolecule #27: Pre-mRNA-splicing factor PRP9
+Macromolecule #28: Pre-mRNA-splicing factor PRP11
+Macromolecule #29: Pre-mRNA-splicing factor PRP21
+Macromolecule #30: U2 small nuclear ribonucleoprotein A'
+Macromolecule #31: Unknown
+Macromolecule #32: U2 small nuclear ribonucleoprotein B''
+Macromolecule #33: RDS3 complex subunit 10
+Macromolecule #34: U6 snRNA-associated Sm-like protein LSm2
+Macromolecule #35: Small nuclear ribonucleoprotein-associated protein B
+Macromolecule #36: Small nuclear ribonucleoprotein Sm D3
+Macromolecule #37: Small nuclear ribonucleoprotein E
+Macromolecule #38: Small nuclear ribonucleoprotein F
+Macromolecule #39: Small nuclear ribonucleoprotein G
+Macromolecule #40: Small nuclear ribonucleoprotein Sm D1
+Macromolecule #41: Small nuclear ribonucleoprotein Sm D2
+Macromolecule #42: U6 snRNA-associated Sm-like protein LSm4
+Macromolecule #43: U6 snRNA-associated Sm-like protein LSm5
+Macromolecule #44: U6 snRNA-associated Sm-like protein LSm6
+Macromolecule #45: GUANOSINE-5'-TRIPHOSPHATE
+Macromolecule #46: ZINC ION
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 1.5 mg/mL | ||||||||||||
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Buffer | pH: 7.9 Component:
Details: Buffer pH: HEPES, 7.9; EDTA, 8.0 | ||||||||||||
Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 15 sec. / Pretreatment - Atmosphere: AIR | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK III Details: Grids were glow-discharged for 15 s before deposition of 3 microliter sample (~1.5 mg mL-1), and subsequently incubated for 2-3.5 s before blotting and vitrification by plunging into liquid ...Details: Grids were glow-discharged for 15 s before deposition of 3 microliter sample (~1.5 mg mL-1), and subsequently incubated for 2-3.5 s before blotting and vitrification by plunging into liquid ethane with a Vitrobot Mark III (FEI) operated at 4 degrees Celsius and 100% humidity.. |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Quantum |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Digitization - Frames/image: 1-20 / Number real images: 5115 / Average exposure time: 16.0 sec. / Average electron dose: 56.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated defocus max: 5.3 µm / Calibrated defocus min: 0.35000000000000003 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal magnification: 81000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Overall B value: 170 |
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Output model | ![]() PDB-5nrl: |