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Yorodumi- EMDB-6340: Electron cryo-microscopy of a BRCA1-RNAPII transcriptional complex -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-6340 | |||||||||
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Title | Electron cryo-microscopy of a BRCA1-RNAPII transcriptional complex | |||||||||
Map data | Reconstruction of BRCA1-RNAPII transcriptional complexes derived from human breast cancer cells | |||||||||
Sample |
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Keywords | transcription / DNA repair / cancer / mRNA / tumor suppressor / ubiquitin ligase | |||||||||
Function / homology | Function and homology information negative regulation of mRNA 3'-end processing / Defective DNA double strand break response due to BRCA1 loss of function / Defective DNA double strand break response due to BARD1 loss of function / : / microfibril binding / BRCA1-BARD1 complex / : / BRCA1-C complex / BRCA1-B complex / BRCA1-A complex ...negative regulation of mRNA 3'-end processing / Defective DNA double strand break response due to BRCA1 loss of function / Defective DNA double strand break response due to BARD1 loss of function / : / microfibril binding / BRCA1-BARD1 complex / : / BRCA1-C complex / BRCA1-B complex / BRCA1-A complex / random inactivation of X chromosome / negative regulation of centriole replication / gamma-tubulin ring complex / negative regulation of intracellular estrogen receptor signaling pathway / nuclear ubiquitin ligase complex / DNA strand resection involved in replication fork processing / chordate embryonic development / negative regulation of fatty acid biosynthetic process / cellular response to indole-3-methanol / homologous recombination / lateral element / tissue homeostasis / XY body / protein K6-linked ubiquitination / regulation of DNA damage checkpoint / regulation of phosphorylation / dosage compensation by inactivation of X chromosome / negative regulation of gene expression via chromosomal CpG island methylation / Impaired BRCA2 binding to PALB2 / : / mitotic G2/M transition checkpoint / negative regulation of protein export from nucleus / postreplication repair / Abortive elongation of HIV-1 transcript in the absence of Tat / DNA repair complex / FGFR2 alternative splicing / RNA polymerase binding / centrosome cycle / Defective homologous recombination repair (HRR) due to BRCA1 loss of function / Defective HDR through Homologous Recombination Repair (HRR) due to PALB2 loss of BRCA1 binding function / Defective HDR through Homologous Recombination Repair (HRR) due to PALB2 loss of BRCA2/RAD51/RAD51C binding function / Homologous DNA Pairing and Strand Exchange / Resolution of D-loop Structures through Synthesis-Dependent Strand Annealing (SDSA) / MicroRNA (miRNA) biogenesis / Resolution of D-loop Structures through Holliday Junction Intermediates / Viral Messenger RNA Synthesis / HDR through Single Strand Annealing (SSA) / Impaired BRCA2 binding to RAD51 / intracellular non-membrane-bounded organelle / Signaling by FGFR2 IIIa TM / response to ionizing radiation / DNA-binding transcription activator activity / RNA Pol II CTD phosphorylation and interaction with CE during HIV infection / RNA Pol II CTD phosphorylation and interaction with CE / Formation of the Early Elongation Complex / Formation of the HIV-1 Early Elongation Complex / mRNA Capping / mRNA Splicing - Minor Pathway / PIWI-interacting RNA (piRNA) biogenesis / HIV Transcription Initiation / RNA Polymerase II HIV Promoter Escape / Transcription of the HIV genome / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Initiation And Promoter Clearance / Transcriptional Regulation by E2F6 / Pausing and recovery of Tat-mediated HIV elongation / Tat-mediated HIV elongation arrest and recovery / mitotic G2 DNA damage checkpoint signaling / Presynaptic phase of homologous DNA pairing and strand exchange / Processing of Capped Intron-Containing Pre-mRNA / RNA polymerase II activity / HIV elongation arrest and recovery / Pausing and recovery of HIV elongation / RNA polymerase II transcribes snRNA genes / negative regulation of cell cycle / positive regulation of vascular endothelial growth factor production / Tat-mediated elongation of the HIV-1 transcript / negative regulation of reactive oxygen species metabolic process / Formation of HIV-1 elongation complex containing HIV-1 Tat / localization / Formation of HIV elongation complex in the absence of HIV Tat / regulation of DNA repair / RNA polymerase II, core complex / protein autoubiquitination / SUMOylation of DNA damage response and repair proteins / ubiquitin ligase complex / RNA Polymerase II Transcription Elongation / Formation of RNA Pol II elongation complex / negative regulation of extrinsic apoptotic signaling pathway via death domain receptors / Maturation of protein E / Maturation of protein E / ER Quality Control Compartment (ERQC) / Myoclonic epilepsy of Lafora / FLT3 signaling by CBL mutants / Prevention of phagosomal-lysosomal fusion / IRAK2 mediated activation of TAK1 complex / Alpha-protein kinase 1 signaling pathway / Glycogen synthesis Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / negative staining / Resolution: 22.0 Å | |||||||||
Authors | Gilmore BL / Winton CE / Demmert AC / Tanner JR / Bowman S / Karageorge V / Patel K / Sheng Z / Kelly DF | |||||||||
Citation | Journal: Nat Struct Mol Biol / Year: 2012 Title: Architecture of the RNA polymerase II preinitiation complex and mechanism of ATP-dependent promoter opening. Authors: Sebastian Grünberg / Linda Warfield / Steven Hahn / Abstract: Yeast RNA polymerase II (Pol II) general transcription factor TFIIE and the TFIIH subunit Ssl2 (yeast ortholog of mammalian XPB) function in the transition of the preinitiation complex (PIC) to the ...Yeast RNA polymerase II (Pol II) general transcription factor TFIIE and the TFIIH subunit Ssl2 (yeast ortholog of mammalian XPB) function in the transition of the preinitiation complex (PIC) to the open complex. We show that the three TFIIE winged-helix (WH) domains form a heterodimer, with the Tfa1 (TFIIEα) WH binding the Pol II clamp and the Tfa2 (TFIIEβ) tandem WH domain encircling promoter DNA that becomes single-stranded in the open complex. Ssl2 lies adjacent to TFIIE, enclosing downstream promoter DNA. Unlike previous proposals, comparison of the PIC and open-complex models strongly suggests that Ssl2 promotes DNA opening by functioning as a double-stranded-DNA translocase, feeding 15 base pairs into the Pol II cleft. Right-handed threading of DNA through the Ssl2 binding groove, combined with the fixed position of upstream promoter DNA, leads to DNA unwinding and the open state. #5: Journal: Journal of Scientific Reports Title: A Molecular Toolkit to Visualize Native Protein Assemblies in the Context of the Human Disease Authors: Gilmore BL / Winton CE / Demmert AC / Tanner JR / Bowman S / Karageorge V / Patel K / Sheng Z / Kelly DF | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_6340.map.gz | 283.4 KB | EMDB map data format | |
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Header (meta data) | emd-6340-v30.xml emd-6340.xml | 23.9 KB 23.9 KB | Display Display | EMDB header |
Images | 400_6340.gif 80_6340.gif | 54.3 KB 4.2 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-6340 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-6340 | HTTPS FTP |
-Related structure data
Similar structure data |
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-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_6340.map.gz / Format: CCP4 / Size: 825.2 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Reconstruction of BRCA1-RNAPII transcriptional complexes derived from human breast cancer cells | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 5.5 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : BRCA1-RNAP II transcriptional assemblies isolated from hereditary...
Entire | Name: BRCA1-RNAP II transcriptional assemblies isolated from hereditary breast cancer cells |
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Components |
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-Supramolecule #1000: BRCA1-RNAP II transcriptional assemblies isolated from hereditary...
Supramolecule | Name: BRCA1-RNAP II transcriptional assemblies isolated from hereditary breast cancer cells type: sample / ID: 1000 Oligomeric state: one BRCA1 molecule binds to one polymerase complex Number unique components: 5 |
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Molecular weight | Theoretical: 800 KDa |
-Macromolecule #1: RNA Polyermase II core
Macromolecule | Name: RNA Polyermase II core / type: protein_or_peptide / ID: 1 / Name.synonym: Pol2 / Number of copies: 1 / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Homo sapiens (human) / synonym: human / Tissue: breast / Cell: tumor / Organelle: nucleus / Location in cell: nucleus |
Molecular weight | Theoretical: 500 KDa |
Sequence | UniProtKB: DNA-directed RNA polymerase II subunit RPB1 |
-Macromolecule #2: BRCA1
Macromolecule | Name: BRCA1 / type: protein_or_peptide / ID: 2 Details: BRCA1 was attached to tunable microchips using antibodies against the BRCA1 C-terminal domain Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Homo sapiens (human) / synonym: human / Tissue: breast / Cell: tumor / Organelle: nucleus / Location in cell: nucleus |
Molecular weight | Theoretical: 200 KDa |
Sequence | UniProtKB: Breast cancer type 1 susceptibility protein |
-Macromolecule #3: BARD1
Macromolecule | Name: BARD1 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Homo sapiens (human) / synonym: human / Tissue: breast / Cell: tumor / Organelle: nucleus / Location in cell: nucleus |
Molecular weight | Theoretical: 87 KDa |
Sequence | UniProtKB: BRCA1-associated RING domain protein 1 |
-Macromolecule #4: Ubiqutin
Macromolecule | Name: Ubiqutin / type: protein_or_peptide / ID: 4 / Number of copies: 2 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Homo sapiens (human) / synonym: human / Tissue: breast / Cell: tumor / Organelle: nucleus / Location in cell: nucleus |
Molecular weight | Theoretical: 8 KDa |
Sequence | UniProtKB: Polyubiquitin-C |
-Macromolecule #5: DNA
Macromolecule | Name: DNA / type: dna / ID: 5 / Classification: DNA / Structure: DOUBLE HELIX / Synthetic?: No |
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Source (natural) | Organism: Homo sapiens (human) / synonym: human |
-Experimental details
-Structure determination
Method | negative staining, cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 / Details: 50 mM HEPES, 150 mM NaCl, 10 mM MgCl2, 10 mM CaCl2 |
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Staining | Type: NEGATIVE Details: Protein complexes were adsorbed to antibody-decorated microchips for 2 minutes and stained with 1% uranyl formate for 1 minute. |
Grid | Details: SiN microchips with TEM windows coated with 25% Ni-NTA functionalized lipid monolayers |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 93 K / Instrument: GATAN CRYOPLUNGE 3 Method: Microchips were blotted for ~8 seconds using one-sided blotting. |
-Electron microscopy
Microscope | FEI TECNAI SPIRIT |
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Electron beam | Acceleration voltage: 120 kV / Electron source: LAB6 |
Electron optics | Calibrated magnification: 54500 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2 mm / Nominal defocus max: -3.0 µm / Nominal defocus min: -1.0 µm / Nominal magnification: 50000 |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
Temperature | Min: 83 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at high magnification. |
Details | low-dose illumination |
Date | Mar 8, 2013 |
Image recording | Category: CCD / Film or detector model: FEI EAGLE (2k x 2k) / Digitization - Sampling interval: 30 µm / Number real images: 50 / Average electron dose: 5 e/Å2 |
Experimental equipment | Model: Tecnai Spirit / Image courtesy: FEI Company |
-Image processing
CTF correction | Details: Each particle |
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Final two d classification | Number classes: 5 |
Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 22.0 Å / Resolution method: OTHER / Software - Name: RELION / Number images used: 22000 |
Details | The particles were selected using an automatic selection program. |
-Atomic model buiding 1
Initial model | PDB ID: Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B / Chain - #2 - Chain ID: C / Chain - #3 - Chain ID: D / Chain - #4 - Chain ID: E / Chain - #5 - Chain ID: F / Chain - #6 - Chain ID: G / Chain - #7 - Chain ID: H / Chain - #8 - Chain ID: I / Chain - #9 - Chain ID: J / Chain - #10 - Chain ID: K / Chain - #11 - Chain ID: L |
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Software | Name: Chimera |
Details | The domains were separately fitted by manual docking using the Chimera software package. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 2
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: Chimera |
Details | The domains were separately fitted by manual docking using the Chimera software package. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 3
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: Chimera |
Details | The domains were separately fitted by manual docking using the Chimera software package. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
-Atomic model buiding 4
Initial model | PDB ID: Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B |
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Software | Name: Chimera |
Details | The domains were separately fitted by manual docking using the Chimera software package. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |