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Open data
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Basic information
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| Title | Cryo-EM Structure of Pre-B+AMPPNP Complex (core part) | |||||||||
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 Keywords | spliceosome / SPLICING | |||||||||
| Function / homology |  Function and homology informationribonucleoprotein complex localization / U4atac snRNP / positive regulation of cytotoxic T cell differentiation / maturation of 5S rRNA / RNA localization / U4atac snRNA binding / R-loop processing / cis assembly of pre-catalytic spliceosome / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / RNA splicing, via transesterification reactions ...ribonucleoprotein complex localization / U4atac snRNP / positive regulation of cytotoxic T cell differentiation / maturation of 5S rRNA / RNA localization / U4atac snRNA binding / R-loop processing / cis assembly of pre-catalytic spliceosome / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / RNA splicing, via transesterification reactions / U2-type catalytic step 1 spliceosome / snRNP binding / U4 snRNA binding / spliceosomal tri-snRNP complex / U2-type precatalytic spliceosome / mRNA cis splicing, via spliceosome / U2-type spliceosomal complex / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / U4 snRNP / U2 snRNP / U2-type prespliceosome / K63-linked polyubiquitin modification-dependent protein binding / precatalytic spliceosome / spliceosomal complex assembly / mRNA Splicing - Minor Pathway / mRNA 3'-splice site recognition / negative regulation of mRNA splicing, via spliceosome / MLL1 complex / spliceosomal tri-snRNP complex assembly / U5 snRNA binding / protein deubiquitination / U5 snRNP / U2 snRNA binding / U6 snRNA binding / pre-mRNA intronic binding / spliceosomal snRNP assembly / ribonucleoprotein complex binding / U1 snRNA binding / Cajal body / U4/U6 x U5 tri-snRNP complex / catalytic step 2 spliceosome / mRNA Splicing - Major Pathway / RNA splicing / response to cocaine / spliceosomal complex / helicase activity / mRNA splicing, via spliceosome / cellular response to xenobiotic stimulus / mRNA processing / osteoblast differentiation / cellular response to tumor necrosis factor / cellular response to lipopolysaccharide / protein-macromolecule adaptor activity / nucleic acid binding / ubiquitinyl hydrolase 1 / RNA helicase activity / hydrolase activity / nuclear speck / cilium / ciliary basal body / RNA helicase / cell division / intracellular membrane-bounded organelle / GTPase activity / mRNA binding / centrosome / chromatin / GTP binding / nucleolus / Golgi apparatus / positive regulation of transcription by RNA polymerase II / ATP hydrolysis activity / RNA binding / extracellular exosome / zinc ion binding / nucleoplasm / ATP binding / identical protein binding / nucleus / membrane / plasma membrane / cytoplasm / cytosol Similarity search - Function  | |||||||||
| Biological species |  Homo sapiens (human) | |||||||||
| Method | single particle reconstruction / cryo EM / Resolution: 4.1 Å | |||||||||
 Authors | Zhang Z / Kumar V / Dybkov O / Will CL / Zhong J / Ludwig S / Urlaub H / Kastner B / Stark H / Luehrmann R | |||||||||
| Funding support |   Germany, 1 items 
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 Citation |  Journal: Nature / Year: 2024Title: Structural insights into the cross-exon to cross-intron spliceosome switch. Authors: Zhenwei Zhang / Vinay Kumar / Olexandr Dybkov / Cindy L Will / Jiayun Zhong / Sebastian E J Ludwig / Henning Urlaub / Berthold Kastner / Holger Stark / Reinhard Lührmann /   ![]() Abstract: Early spliceosome assembly can occur through an intron-defined pathway, whereby U1 and U2 small nuclear ribonucleoprotein particles (snRNPs) assemble across the intron. Alternatively, it can occur ...Early spliceosome assembly can occur through an intron-defined pathway, whereby U1 and U2 small nuclear ribonucleoprotein particles (snRNPs) assemble across the intron. Alternatively, it can occur through an exon-defined pathway, whereby U2 binds the branch site located upstream of the defined exon and U1 snRNP interacts with the 5' splice site located directly downstream of it. The U4/U6.U5 tri-snRNP subsequently binds to produce a cross-intron (CI) or cross-exon (CE) pre-B complex, which is then converted to the spliceosomal B complex. Exon definition promotes the splicing of upstream introns and plays a key part in alternative splicing regulation. However, the three-dimensional structure of exon-defined spliceosomal complexes and the molecular mechanism of the conversion from a CE-organized to a CI-organized spliceosome, a pre-requisite for splicing catalysis, remain poorly understood. Here cryo-electron microscopy analyses of human CE pre-B complex and B-like complexes reveal extensive structural similarities with their CI counterparts. The results indicate that the CE and CI spliceosome assembly pathways converge already at the pre-B stage. Add-back experiments using purified CE pre-B complexes, coupled with cryo-electron microscopy, elucidate the order of the extensive remodelling events that accompany the formation of B complexes and B-like complexes. The molecular triggers and roles of B-specific proteins in these rearrangements are also identified. We show that CE pre-B complexes can productively bind in trans to a U1 snRNP-bound 5' splice site. Together, our studies provide new mechanistic insights into the CE to CI switch during spliceosome assembly and its effect on pre-mRNA splice site pairing at this stage.  | |||||||||
| History | 
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Structure visualization
| Supplemental images | 
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Downloads & links
-EMDB archive
| Map data |  emd_18545.map.gz | 387.7 MB |  EMDB map data format | |
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| Header (meta data) |  emd-18545-v30.xml emd-18545.xml | 37.5 KB 37.5 KB  | Display Display  |  EMDB header | 
| FSC (resolution estimation) |  emd_18545_fsc.xml | 16.9 KB | Display |  FSC data file | 
| Images |  emd_18545.png | 41.1 KB | ||
| Filedesc metadata |  emd-18545.cif.gz | 13.6 KB | ||
| Others |  emd_18545_half_map_1.map.gz emd_18545_half_map_2.map.gz | 337.1 MB 337 MB  | ||
| Archive directory |  http://ftp.pdbj.org/pub/emdb/structures/EMD-18545 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-18545 | HTTPS FTP  | 
-Validation report
| Summary document |  emd_18545_validation.pdf.gz | 930.2 KB | Display |  EMDB validaton report | 
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| Full document |  emd_18545_full_validation.pdf.gz | 929.7 KB | Display | |
| Data in XML |  emd_18545_validation.xml.gz | 24.5 KB | Display | |
| Data in CIF |  emd_18545_validation.cif.gz | 32.7 KB | Display | |
| Arichive directory |  https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-18545 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-18545 | HTTPS FTP  | 
-Related structure data
| Related structure data | ![]() 8qp9MC ![]() 8qozC ![]() 8qp8C ![]() 8qpaC ![]() 8qpbC ![]() 8qpeC ![]() 8qpkC ![]() 8qxdC ![]() 8qzsC ![]() 8r08C ![]() 8r09C ![]() 8r0aC ![]() 8r0bC ![]() 8rm5C M: atomic model generated by this map C: citing same article (  | 
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| Similar structure data | Similarity search - Function & homology  F&H Search | 
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Links
| EMDB pages |  EMDB (EBI/PDBe) /  EMDataResource | 
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| Related items in Molecule of the Month | 
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Map
| File |  Download / File: emd_18545.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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| Projections & slices | Image control
 
 Images are generated by Spider.  | ||||||||||||||||||||||||||||||||||||
| Voxel size | X=Y=Z: 1.35 Å | ||||||||||||||||||||||||||||||||||||
| Density | 
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| Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
| Details | EMDB XML: 
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-Supplemental data
-Half map: #2
| File | emd_18545_half_map_1.map | ||||||||||||
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| Projections & Slices | 
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| Density Histograms | 
-Half map: #1
| File | emd_18545_half_map_2.map | ||||||||||||
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| Projections & Slices | 
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| Density Histograms | 
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Sample components
+Entire : human spliceosomal pre-B+AMPPNP complex
+Supramolecule #1: human spliceosomal pre-B+AMPPNP complex
+Macromolecule #1: Splicing factor 3A subunit 1
+Macromolecule #2: U5 small nuclear ribonucleoprotein 200 kDa helicase
+Macromolecule #3: 116 kDa U5 small nuclear ribonucleoprotein component
+Macromolecule #4: Thioredoxin-like protein 4A
+Macromolecule #5: U4/U6 small nuclear ribonucleoprotein Prp3
+Macromolecule #6: U4/U6 small nuclear ribonucleoprotein Prp31
+Macromolecule #7: RNA-binding protein 42
+Macromolecule #8: U4/U6.U5 tri-snRNP-associated protein 1
+Macromolecule #9: Ubiquitin carboxyl-terminal hydrolase 39
+Macromolecule #10: U4/U6.U5 small nuclear ribonucleoprotein 27 kDa protein
+Macromolecule #11: Probable ATP-dependent RNA helicase DDX23
+Macromolecule #15: Pre-mRNA-processing-splicing factor 8
+Macromolecule #16: Pre-mRNA-processing factor 6
+Macromolecule #12: U6 snRNA
+Macromolecule #13: U5 snRNA
+Macromolecule #14: U4 snRNA
+Macromolecule #17: INOSITOL HEXAKISPHOSPHATE
-Experimental details
-Structure determination
| Method | cryo EM | 
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 Processing | single particle reconstruction | 
| Aggregation state | particle | 
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Sample preparation
| Buffer | pH: 7.9 | 
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| Vitrification | Cryogen name: ETHANE | 
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Electron microscopy
| Microscope | FEI TITAN KRIOS | 
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| Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Average electron dose: 45.0 e/Å2 | 
| Electron beam | Acceleration voltage: 300 kV / Electron source:  FIELD EMISSION GUN | 
| Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.0 µm / Nominal defocus min: 1.5 µm | 
| Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company  | 
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About Yorodumi




Keywords
Homo sapiens (human)
Authors
Germany, 1 items 
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Processing
FIELD EMISSION GUN

