+Open data
-Basic information
Entry | Database: PDB / ID: 7os2 | ||||||
---|---|---|---|---|---|---|---|
Title | Cryo-EM structure of Brr2 in complex with Jab1/MPN and C9ORF78 | ||||||
Components |
| ||||||
Keywords | SPLICING / mRNA Splicing / Spliceosomal Assembly / Splicing Regulation / Brr2 Helicase | ||||||
Function / homology | Function and homology information regulation of homologous chromosome segregation / cis assembly of pre-catalytic spliceosome / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / U2-type catalytic step 1 spliceosome / RNA splicing, via transesterification reactions / mRNA cis splicing, via spliceosome / U2-type precatalytic spliceosome / U2-type catalytic step 2 spliceosome / K63-linked polyubiquitin modification-dependent protein binding / mRNA Splicing - Minor Pathway ...regulation of homologous chromosome segregation / cis assembly of pre-catalytic spliceosome / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / U2-type catalytic step 1 spliceosome / RNA splicing, via transesterification reactions / mRNA cis splicing, via spliceosome / U2-type precatalytic spliceosome / U2-type catalytic step 2 spliceosome / K63-linked polyubiquitin modification-dependent protein binding / mRNA Splicing - Minor Pathway / chromosome, centromeric region / spliceosomal tri-snRNP complex assembly / U5 snRNA binding / U5 snRNP / U2 snRNA binding / U6 snRNA binding / pre-mRNA intronic binding / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / catalytic step 2 spliceosome / mRNA Splicing - Major Pathway / RNA splicing / helicase activity / chromosome segregation / spliceosomal complex / mRNA processing / mRNA splicing, via spliceosome / osteoblast differentiation / cellular response to tumor necrosis factor / cellular response to lipopolysaccharide / RNA helicase activity / RNA helicase / nuclear speck / ATP hydrolysis activity / RNA binding / nucleoplasm / ATP binding / identical protein binding / membrane / nucleus / cytosol Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.76 Å | ||||||
Authors | Bergfort, A. / Hilal, T. / Weber, G. / Wahl, M.C. | ||||||
Funding support | Germany, 1items
| ||||||
Citation | Journal: Nucleic Acids Res / Year: 2022 Title: The intrinsically disordered TSSC4 protein acts as a helicase inhibitor, placeholder and multi-interaction coordinator during snRNP assembly and recycling. Authors: Alexandra Bergfort / Tarek Hilal / Benno Kuropka / İbrahim Avşar Ilik / Gert Weber / Tuğçe Aktaş / Christian Freund / Markus C Wahl / Abstract: Biogenesis of spliceosomal small nuclear ribonucleoproteins (snRNPs) and their recycling after splicing require numerous assembly/recycling factors whose modes of action are often poorly understood. ...Biogenesis of spliceosomal small nuclear ribonucleoproteins (snRNPs) and their recycling after splicing require numerous assembly/recycling factors whose modes of action are often poorly understood. The intrinsically disordered TSSC4 protein has been identified as a nuclear-localized U5 snRNP and U4/U6-U5 tri-snRNP assembly/recycling factor, but how TSSC4's intrinsic disorder supports TSSC4 functions remains unknown. Using diverse interaction assays and cryogenic electron microscopy-based structural analysis, we show that TSSC4 employs four conserved, non-contiguous regions to bind the PRPF8 Jab1/MPN domain and the SNRNP200 helicase at functionally important sites. It thereby inhibits SNRNP200 helicase activity, spatially aligns the proteins, coordinates formation of a U5 sub-module and transiently blocks premature interaction of SNRNP200 with at least three other spliceosomal factors. Guided by the structure, we designed a TSSC4 variant that lacks stable binding to the PRPF8 Jab1/MPN domain or SNRNP200 in vitro. Comparative immunoprecipitation/mass spectrometry from HEK293 nuclear extract revealed distinct interaction profiles of wild type TSSC4 and the variant deficient in PRPF8/SNRNP200 binding with snRNP proteins, other spliceosomal proteins as well as snRNP assembly/recycling factors and chaperones. Our findings elucidate molecular strategies employed by an intrinsically disordered protein to promote snRNP assembly, and suggest multiple TSSC4-dependent stages during snRNP assembly/recycling. #1: Journal: Acta Crystallogr D Struct Biol / Year: 2019 Title: Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix. Authors: Liebschner, D. / Afonine, P.V. / Baker, M.L. / Bunkoczi, G. / Chen, V.B. / Croll, T.I. / Hintze, B. / Hung, L.W. / Jain, S. / McCoy, A.J. / Moriarty, N.W. / Oeffner, R.D. / Poon, B.K. / ...Authors: Liebschner, D. / Afonine, P.V. / Baker, M.L. / Bunkoczi, G. / Chen, V.B. / Croll, T.I. / Hintze, B. / Hung, L.W. / Jain, S. / McCoy, A.J. / Moriarty, N.W. / Oeffner, R.D. / Poon, B.K. / Prisant, M.G. / Read, R.J. / Richardson, J.S. / Richardson, D.C. / Sammito, M.D. / Sobolev, O.V. / Stockwell, D.H. / Terwilliger, T.C. / Urzhumtsev, A.G. / Videau, L.L. / Williams, C.J. / Adams, P.D. | ||||||
History |
|
-Structure visualization
Movie |
Movie viewer |
---|---|
Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7os2.cif.gz | 390.4 KB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb7os2.ent.gz | 298 KB | Display | PDB format |
PDBx/mmJSON format | 7os2.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7os2_validation.pdf.gz | 810.3 KB | Display | wwPDB validaton report |
---|---|---|---|---|
Full document | 7os2_full_validation.pdf.gz | 831.4 KB | Display | |
Data in XML | 7os2_validation.xml.gz | 62.1 KB | Display | |
Data in CIF | 7os2_validation.cif.gz | 94.5 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/os/7os2 ftp://data.pdbj.org/pub/pdb/validation_reports/os/7os2 | HTTPS FTP |
-Related structure data
Related structure data | 13046MC 7os1C 7px3C M: map data used to model this data C: citing same article (ref.) |
---|---|
Similar structure data |
-Links
-Assembly
Deposited unit |
|
---|---|
1 |
|
-Components
#1: Protein | Mass: 198785.797 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: O75643, RNA helicase |
---|---|
#2: Protein | Mass: 34081.516 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: C9orf78 / Production host: Escherichia coli (E. coli) / References: UniProt: Q9NZ63 |
#3: Protein | Mass: 31860.984 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: PRPF8 / Production host: Escherichia coli (E. coli) / References: UniProt: Q6P2Q9 |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component |
| ||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Molecular weight |
| ||||||||||||||||||||||||||||||
Source (natural) |
| ||||||||||||||||||||||||||||||
Source (recombinant) |
| ||||||||||||||||||||||||||||||
Buffer solution | pH: 7.6 | ||||||||||||||||||||||||||||||
Specimen | Conc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 283 K |
-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: 120000 X / Nominal defocus max: 1800 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: ZEMLIN TABLEAU |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 40 sec. / Electron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 5160 |
-Processing
Software |
| |||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
EM software |
| |||||||||||||||||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 936716 | |||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.76 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 370493 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 102 / Protocol: OTHER / Space: REAL / Target criteria: CC, R.m.s.d. Bonds / Angles | |||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | 3D fitting-ID: 1 / Accession code: 4KIT / Initial refinement model-ID: 1 / PDB-ID: 4KIT / Source name: PDB / Type: experimental model
| |||||||||||||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | |||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 101.73 Å2 | |||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
|