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Yorodumi- PDB-6urg: Cryo-EM structure of human CPSF160-WDR33-CPSF30-CPSF100 PIM complex -
+Open data
-Basic information
Entry | Database: PDB / ID: 6urg | ||||||
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Title | Cryo-EM structure of human CPSF160-WDR33-CPSF30-CPSF100 PIM complex | ||||||
Components |
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Keywords | PROTEIN BINDING / pre-mRNA 3'-end processing / mammalin cleavage factor(mCF) / CPSF100 / mPSF | ||||||
Function / homology | Function and homology information co-transcriptional RNA 3'-end processing, cleavage and polyadenylation pathway / Inhibition of Host mRNA Processing and RNA Silencing / mRNA 3'-end processing by stem-loop binding and cleavage / Processing of Intronless Pre-mRNAs / mRNA cleavage and polyadenylation specificity factor complex / collagen trimer / mRNA 3'-UTR AU-rich region binding / Transport of Mature mRNA Derived from an Intronless Transcript / mRNA 3'-end processing / mRNA 3'-end processing ...co-transcriptional RNA 3'-end processing, cleavage and polyadenylation pathway / Inhibition of Host mRNA Processing and RNA Silencing / mRNA 3'-end processing by stem-loop binding and cleavage / Processing of Intronless Pre-mRNAs / mRNA cleavage and polyadenylation specificity factor complex / collagen trimer / mRNA 3'-UTR AU-rich region binding / Transport of Mature mRNA Derived from an Intronless Transcript / mRNA 3'-end processing / mRNA 3'-end processing / tRNA processing in the nucleus / postreplication repair / RNA Polymerase II Transcription Termination / Processing of Capped Intron-Containing Pre-mRNA / fibrillar center / mRNA processing / sequence-specific double-stranded DNA binding / spermatogenesis / postsynapse / intracellular membrane-bounded organelle / glutamatergic synapse / enzyme binding / RNA binding / zinc ion binding / nucleoplasm / membrane / nucleus Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å | ||||||
Authors | Sun, Y. / Zhang, Y. / Walz, T. / Tong, L. | ||||||
Funding support | United States, 1items
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Citation | Journal: Mol Cell / Year: 2020 Title: Structural Insights into the Human Pre-mRNA 3'-End Processing Machinery. Authors: Yixiao Zhang / Yadong Sun / Yongsheng Shi / Thomas Walz / Liang Tong / Abstract: The mammalian pre-mRNA 3'-end-processing machinery consists of cleavage and polyadenylation specificity factor (CPSF), cleavage stimulation factor (CstF), and other proteins, but the overall ...The mammalian pre-mRNA 3'-end-processing machinery consists of cleavage and polyadenylation specificity factor (CPSF), cleavage stimulation factor (CstF), and other proteins, but the overall architecture of this machinery remains unclear. CPSF contains two functionally distinct modules: a cleavage factor (mCF) and a polyadenylation specificity factor (mPSF). Here, we have produced recombinant human CPSF and CstF and examined these factors by electron microscopy (EM). We find that mPSF is the organizational core of the machinery, while the conformations of mCF and CstF and the position of mCF relative to mPSF are highly variable. We have identified by cryo-EM a segment in CPSF100 that tethers mCF to mPSF, and we have named it the PSF interaction motif (PIM). Mutations in the PIM can abolish CPSF formation, indicating that it is a crucial contact in CPSF. We have also obtained reconstructions of mCF and CstF77 by cryo-EM, assembled around the mPSF core. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6urg.cif.gz | 333.5 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6urg.ent.gz | 244.9 KB | Display | PDB format |
PDBx/mmJSON format | 6urg.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6urg_validation.pdf.gz | 802.9 KB | Display | wwPDB validaton report |
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Full document | 6urg_full_validation.pdf.gz | 820.3 KB | Display | |
Data in XML | 6urg_validation.xml.gz | 48.5 KB | Display | |
Data in CIF | 6urg_validation.cif.gz | 72.4 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ur/6urg ftp://data.pdbj.org/pub/pdb/validation_reports/ur/6urg | HTTPS FTP |
-Related structure data
Related structure data | 20860MC 6uroC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 161074.234 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: CPSF1, CPSF160 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q10570 |
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#2: Protein | Mass: 67546.812 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: WDR33, WDC146 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9C0J8 |
#3: Protein | Mass: 28417.883 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: CPSF4, CPSF30, NAR, NEB1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: O95639 |
#4: Protein | Mass: 88597.734 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: CPSF2, CPSF100, KIAA1367 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9P2I0 |
#5: Chemical | ChemComp-ZN / |
Has ligand of interest | N |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: The complex of human mPSF-CPSF100 PIM / Type: COMPLEX / Entity ID: #1-#4 / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Trichoplusia ni (cabbage looper) |
Buffer solution | pH: 8 / Details: 25 mM Tris-HCl, pH 8.0, 150 mM NaCl, 5 mM DTT |
Buffer component | Conc.: 150 mM / Name: sodium chloride / Formula: NaCl |
Specimen | Conc.: 0.25 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: unspecified |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-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: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 225000 X / Calibrated magnification: 46729 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1200 nm / Calibrated defocus min: 900 nm / Calibrated defocus max: 2800 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 10 sec. / Electron dose: 71 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
Software | Name: PHENIX / Version: 1.12_2829: / Classification: refinement | ||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 859796 / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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