+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-3440 | |||||||||
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Title | Structure of the Saccharomyces cerevisiae TREX-2 complex | |||||||||
Map data | Postprocessed sharpened map, masked | |||||||||
Sample |
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Keywords | mRNA export | |||||||||
Function / homology | Function and homology information actin filament-based process / cellular bud site selection / SAGA complex localization to transcription regulatory region / transcription export complex 2 / post-transcriptional tethering of RNA polymerase II gene DNA at nuclear periphery / nuclear pore cytoplasmic filaments / nuclear mRNA surveillance / maintenance of DNA trinucleotide repeats / filamentous growth / mRNA 3'-end processing ...actin filament-based process / cellular bud site selection / SAGA complex localization to transcription regulatory region / transcription export complex 2 / post-transcriptional tethering of RNA polymerase II gene DNA at nuclear periphery / nuclear pore cytoplasmic filaments / nuclear mRNA surveillance / maintenance of DNA trinucleotide repeats / filamentous growth / mRNA 3'-end processing / proteasome regulatory particle, lid subcomplex / poly(A)+ mRNA export from nucleus / proteasome storage granule / transcription-coupled nucleotide-excision repair / proteasome assembly / mRNA export from nucleus / protein folding chaperone / protein export from nucleus / proteasome complex / transcription elongation by RNA polymerase II / double-strand break repair via homologous recombination / ribosomal small subunit biogenesis / mitotic cell cycle / nuclear envelope / ubiquitin-dependent protein catabolic process / double-stranded DNA binding / proteasome-mediated ubiquitin-dependent protein catabolic process / molecular adaptor activity / regulation of cell cycle / protein-containing complex binding / positive regulation of transcription by RNA polymerase II / RNA binding / nucleus / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Saccharomyces cerevisae | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 5.3 Å | |||||||||
Authors | Aibara S / Bai XC / Stewart M | |||||||||
Citation | Journal: J Struct Biol / Year: 2016 Title: The Sac3 TPR-like region in the Saccharomyces cerevisiae TREX-2 complex is more extensive but independent of the CID region. Authors: Shintaro Aibara / Xiao-Chen Bai / Murray Stewart / Abstract: Transcription-export complex 2 (TREX-2 complex) facilitates the localization of actively transcribing genes to the nuclear periphery and also functions to contribute to the generation of export- ...Transcription-export complex 2 (TREX-2 complex) facilitates the localization of actively transcribing genes to the nuclear periphery and also functions to contribute to the generation of export-competent mRNPs through interactions with the general mRNA nuclear export factor Mex67:Mtr2. The TREX-2 complex is based on a Sac3 scaffold to which Thp1, Sem1, Cdc31, and Sus1 bind. TREX-2 can be subdivided into two modules: one, in which Thp1 and Sem1 bind to the Sac3(M) region (residues ∼100-551), and the other in which Cdc31 and two Sus1 chains bind to the Sac3(CID) region (residues ∼710-805). Complementary structural analyses using X-ray crystallography, electron microscopy, and small-angle X-ray scattering of the Saccharomyces cerevisiae TREX-2 complex, expressed using Baculovirus-infected Sf9 cells, have indicated that the TPR-like repeats of the Sac3(M) region extend considerably further towards the N-terminus than previously thought, and also indicate that this region and Sac3(CID):Sus1:Cdc31 region of the S. cerevisiae complex are structurally independent. Although the density visible accounted for only ∼100kDa, a 5.3Å resolution cryo-EM reconstruction was obtained of the M-region of TREX-2 that showed an additional three putative α-helices extending towards the Sac3 N-terminus and these helices were also seen in a 4.9Å resolution structure obtained by X-ray crystallography. SUMMARY STATEMENT: We describe the expression, purification and structural characterization of the S. cerevisiae TREX-2 complex and demonstrate that the Sac3 TPR-like repeats are more extensive than ...SUMMARY STATEMENT: We describe the expression, purification and structural characterization of the S. cerevisiae TREX-2 complex and demonstrate that the Sac3 TPR-like repeats are more extensive than previously thought and that the M- and CID-regions do not appear to have a defined spatial orientation. | |||||||||
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_3440.map.gz | 1021.9 KB | EMDB map data format | |
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Header (meta data) | emd-3440-v30.xml emd-3440.xml | 13.3 KB 13.3 KB | Display Display | EMDB header |
Images | emd-3440.png | 163.7 KB | ||
Masks | emd_3440_msk_1.map | 10.5 MB | Mask map | |
Others | run1_half1_class001_unfil.map run1_half2_class001_unfil_1.map | 10.5 MB 10.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-3440 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-3440 | HTTPS FTP |
-Related structure data
Related structure data | 5g5pMC 5l3tC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_3440.map.gz / Format: CCP4 / Size: 10.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Postprocessed sharpened map, masked | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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
-Segmentation: Automask used for postprocessing
Annotation | Automask used for postprocessing | ||||||||||||
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File | emd_3440_msk_1.map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Supplemental map: run1 half1 class001 unfil.map
File | run1_half1_class001_unfil.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Supplemental map: run1 half2 class001 unfil 1.map
File | run1_half2_class001_unfil_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Sac3 in complex with Thp1 and Sem1
Entire | Name: Sac3 in complex with Thp1 and Sem1 |
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Components |
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-Supramolecule #1000: Sac3 in complex with Thp1 and Sem1
Supramolecule | Name: Sac3 in complex with Thp1 and Sem1 / type: sample / ID: 1000 / Details: Sample is monodisperse / Oligomeric state: One heterotrimer / Number unique components: 3 |
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Molecular weight | Experimental: 190 KDa / Theoretical: 190 KDa / Method: SAXS |
-Macromolecule #1: Sac3
Macromolecule | Name: Sac3 / type: protein_or_peptide / ID: 1 / Details: Construct had residues 1-805 / Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Saccharomyces cerevisae / synonym: Yeast |
Molecular weight | Theoretical: 149.5916 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) / Recombinant cell: sf9 / Recombinant plasmid: pAceBacRZ |
Sequence | UniProtKB: Nuclear mRNA export protein SAC3 |
-Macromolecule #2: Thp1
Macromolecule | Name: Thp1 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Saccharomyces cerevisae / synonym: Yeast |
Molecular weight | Theoretical: 52.6857 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) / Recombinant cell: sf9 / Recombinant plasmid: pAceBacRZ |
Sequence | UniProtKB: Nuclear mRNA export protein THP1 |
-Macromolecule #3: Sem1
Macromolecule | Name: Sem1 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Oligomeric state: Monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Saccharomyces cerevisae / synonym: Yeast |
Molecular weight | Theoretical: 10.372 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) / Recombinant cell: sf9 / Recombinant plasmid: pAceBacRZ |
Sequence | UniProtKB: 26S proteasome complex subunit SEM1 |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.015 mg/mL |
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Buffer | pH: 8 / Details: 20 mM HEPES, 300 mM NaCl, 5 mM DTT |
Grid | Details: Graphene oxide treated 300 mesh gold quantifoil R2/2 grids. |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 85 K / Instrument: HOMEMADE PLUNGER / Method: Blot for 7 seconds before plunging from one side |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 35714 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 81000 |
Specialist optics | Energy filter - Name: Gatan Quantum / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Temperature | Min: 80 K / Max: 90 K / Average: 85 K |
Date | Nov 3, 2015 |
Image recording | Category: CCD / Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Average electron dose: 40 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
CTF correction | Details: Each particle |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 5.3 Å / Resolution method: OTHER / Software - Name: Relion / Number images used: 81559 |
Details | The particles were processed using relion. |