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Yorodumi- EMDB-2875: Cryo electron microscopy of actively translating human polysomes ... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-2875 | |||||||||
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Title | Cryo electron microscopy of actively translating human polysomes (POST state). | |||||||||
Map data | POST state of human polysomes | |||||||||
Sample |
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Keywords | mammalian ribosome / translation / polysome / cryo electron microscopy / elongation cycle | |||||||||
Function / homology | Function and homology information positive regulation of cysteine-type endopeptidase activity involved in execution phase of apoptosis / negative regulation of endoplasmic reticulum unfolded protein response / embryonic brain development / oxidized pyrimidine DNA binding / response to TNF agonist / positive regulation of base-excision repair / eukaryotic 80S initiation complex / protein tyrosine kinase inhibitor activity / negative regulation of protein neddylation / translation at presynapse ...positive regulation of cysteine-type endopeptidase activity involved in execution phase of apoptosis / negative regulation of endoplasmic reticulum unfolded protein response / embryonic brain development / oxidized pyrimidine DNA binding / response to TNF agonist / positive regulation of base-excision repair / eukaryotic 80S initiation complex / protein tyrosine kinase inhibitor activity / negative regulation of protein neddylation / translation at presynapse / positive regulation of intrinsic apoptotic signaling pathway in response to DNA damage / positive regulation of respiratory burst involved in inflammatory response / positive regulation of gastrulation / axial mesoderm development / regulation of G1 to G0 transition / negative regulation of formation of translation preinitiation complex / IRE1-RACK1-PP2A complex / nucleolus organization / ribosomal protein import into nucleus / positive regulation of intrinsic apoptotic signaling pathway in response to DNA damage by p53 class mediator / regulation of translation involved in cellular response to UV / response to extracellular stimulus / positive regulation of endodeoxyribonuclease activity / positive regulation of Golgi to plasma membrane protein transport / exit from mitosis / protein-DNA complex disassembly / TNFR1-mediated ceramide production / 90S preribosome assembly / positive regulation of DNA damage response, signal transduction by p53 class mediator resulting in transcription of p21 class mediator / negative regulation of DNA repair / negative regulation of RNA splicing / laminin receptor activity / optic nerve development / TORC2 complex binding / oxidized purine DNA binding / negative regulation of intrinsic apoptotic signaling pathway in response to hydrogen peroxide / supercoiled DNA binding / GAIT complex / G1 to G0 transition / neural crest cell differentiation / retinal ganglion cell axon guidance / NF-kappaB complex / rRNA modification in the nucleus and cytosol / negative regulation of phagocytosis / middle ear morphogenesis / ubiquitin-like protein conjugating enzyme binding / regulation of establishment of cell polarity / positive regulation of ubiquitin-protein transferase activity / Formation of the ternary complex, and subsequently, the 43S complex / erythrocyte homeostasis / cytoplasmic side of rough endoplasmic reticulum membrane / A band / positive regulation of signal transduction by p53 class mediator / ubiquitin ligase inhibitor activity / alpha-beta T cell differentiation / pigmentation / protein kinase A binding / negative regulation of ubiquitin protein ligase activity / Ribosomal scanning and start codon recognition / ion channel inhibitor activity / Translation initiation complex formation / phagocytic cup / positive regulation of mitochondrial depolarization / response to aldosterone / negative regulation of Wnt signaling pathway / homeostatic process / positive regulation of T cell receptor signaling pathway / lung morphogenesis / macrophage chemotaxis / positive regulation of activated T cell proliferation / fibroblast growth factor binding / regulation of cell division / SARS-CoV-1 modulates host translation machinery / iron-sulfur cluster binding / Protein hydroxylation / TOR signaling / BH3 domain binding / mTORC1-mediated signalling / endonucleolytic cleavage to generate mature 3'-end of SSU-rRNA from (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / Peptide chain elongation / Selenocysteine synthesis / protein-RNA complex assembly / monocyte chemotaxis / cysteine-type endopeptidase activator activity involved in apoptotic process / Formation of a pool of free 40S subunits / ribosomal small subunit export from nucleus / positive regulation of cyclic-nucleotide phosphodiesterase activity / Eukaryotic Translation Termination / blastocyst development / Response of EIF2AK4 (GCN2) to amino acid deficiency / translation regulator activity / SRP-dependent cotranslational protein targeting to membrane / positive regulation of intrinsic apoptotic signaling pathway by p53 class mediator / Viral mRNA Translation / protein localization to nucleus / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / cellular response to actinomycin D / GTP hydrolysis and joining of the 60S ribosomal subunit / negative regulation of proteasomal ubiquitin-dependent protein catabolic process / negative regulation of respiratory burst involved in inflammatory response Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.5 Å | |||||||||
Authors | Behrmann E / Loerke J / Budkevich TV / Yamamoto K / Schmidt A / Penczek PA / Vos MR / Burger J / Mielke T / Scheerer P / Spahn CMT | |||||||||
Citation | Journal: Cell / Year: 2015 Title: Structural snapshots of actively translating human ribosomes. Authors: Elmar Behrmann / Justus Loerke / Tatyana V Budkevich / Kaori Yamamoto / Andrea Schmidt / Pawel A Penczek / Matthijn R Vos / Jörg Bürger / Thorsten Mielke / Patrick Scheerer / Christian M T Spahn / Abstract: Macromolecular machines, such as the ribosome, undergo large-scale conformational changes during their functional cycles. Although their mode of action is often compared to that of mechanical ...Macromolecular machines, such as the ribosome, undergo large-scale conformational changes during their functional cycles. Although their mode of action is often compared to that of mechanical machines, a crucial difference is that, at the molecular dimension, thermodynamic effects dominate functional cycles, with proteins fluctuating stochastically between functional states defined by energetic minima on an energy landscape. Here, we have used cryo-electron microscopy to image ex-vivo-derived human polysomes as a source of actively translating ribosomes. Multiparticle refinement and 3D variability analysis allowed us to visualize a variety of native translation intermediates. Significantly populated states include not only elongation cycle intermediates in pre- and post-translocational states, but also eEF1A-containing decoding and termination/recycling complexes. Focusing on the post-translocational state, we extended this assessment to the single-residue level, uncovering striking details of ribosome-ligand interactions and identifying both static and functionally important dynamic elements. | |||||||||
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_2875.map.gz | 152.8 MB | EMDB map data format | |
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Header (meta data) | emd-2875-v30.xml emd-2875.xml | 10.9 KB 10.9 KB | Display Display | EMDB header |
Images | post.tif | 3.1 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-2875 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-2875 | HTTPS FTP |
-Related structure data
Related structure data | 5aj0MUC 2902C 2903C 2904C 2905C 2906C 2907C 2908C 2909C 2910C 2911C M: atomic model generated by this map U: unfit; in different coordinate system*YM 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_2875.map.gz / Format: CCP4 / Size: 238.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | POST state of human polysomes | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.945 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 : Native ribosomal complex from polysomes - POST state
Entire | Name: Native ribosomal complex from polysomes - POST state |
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Components |
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-Supramolecule #1000: Native ribosomal complex from polysomes - POST state
Supramolecule | Name: Native ribosomal complex from polysomes - POST state / type: sample / ID: 1000 / Details: The sample was monodisperse / Number unique components: 1 |
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Molecular weight | Experimental: 4.5 MDa / Theoretical: 4.5 MDa |
-Supramolecule #1: 80S ribosomal complex
Supramolecule | Name: 80S ribosomal complex / type: complex / ID: 1 / Name.synonym: 80S ribosome Details: The sample was isolated as polysomes from the cell. Structural analysis shows that the complex contains 2 tRNAs and mRNA. Recombinant expression: No / Ribosome-details: ribosome-eukaryote: ALL |
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Source (natural) | Organism: Homo sapiens (human) / Strain: HEK 293T / synonym: Human / Tissue: Kidney / Cell: HEK 293T / Location in cell: Cytoplasm |
Molecular weight | Experimental: 4.5 MDa / Theoretical: 4.5 MDa |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 3.5 mg/mL |
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Buffer | pH: 7.5 Details: 20 mM Hepes-KOH, pH 7.5, 100 mM KCl, 1.5 mM MgCl2, 0.5 mM spermidine, 0.04 mM spermine, 1 mM DTT |
Grid | Details: Quantifoil grids with additional continuous carbon support |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 93 K / Instrument: FEI VITROBOT MARK II / Method: blot for 2-4 seconds before plunging |
-Electron microscopy #1
Microscope | FEI POLARA 300 |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 205000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 115000 |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
Microscopy ID | 1 |
Details | Data was collected automatically with Leginon |
Date | Aug 20, 2012 |
Image recording | Category: CCD / Film or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Number real images: 51282 / Average electron dose: 20 e/Å2 |
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |
-Electron microscopy #2
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 172000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 96000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Microscopy ID | 2 |
Date | Nov 29, 2012 |
Image recording | Category: CCD / Film or detector model: FEI FALCON II (4k x 4k) / Number real images: 51282 / Average electron dose: 20 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
CTF correction | Details: Each micrograph |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 3.5 Å / Resolution method: OTHER / Software - Name: Signature, CTFFind3, Spider, SPARX Details: Maps were calculated from 2 datasets using SSNR-weighted combination. Number images used: 313321 |