|Entry||Database: PDB / ID: 6hyd|
|Title||Rea1 Wild type ADP state (tail part)|
|Keywords||MOTOR PROTEIN / Rea1 / Mdn1 / Midasin / AAA+ protein / ribosome maturation / molecular machine|
|Function / homology||von Willebrand factor, type A / ATPase, dynein-related, AAA domain / VWFA domain profile. / AAA domain (dynein-related subfamily) / von Willebrand factor A-like domain superfamily / P-loop containing nucleoside triphosphate hydrolase / Sigma-54 interaction domain, ATP-binding site 1 / Midasin / AAA+ ATPase domain / regulation of ribosomal subunit export from nucleus ...von Willebrand factor, type A / ATPase, dynein-related, AAA domain / VWFA domain profile. / AAA domain (dynein-related subfamily) / von Willebrand factor A-like domain superfamily / P-loop containing nucleoside triphosphate hydrolase / Sigma-54 interaction domain, ATP-binding site 1 / Midasin / AAA+ ATPase domain / regulation of ribosomal subunit export from nucleus / protein-containing complex remodeling / rRNA processing / ribosomal large subunit assembly / ATPase activity / nucleolus / mitochondrion / nucleoplasm / ATP binding / nucleus / Midasin|
Function and homology information
|Specimen source||Saccharomyces cerevisiae (baker's yeast)|
|Method||ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / 3.9 Å resolution|
|Authors||Sosnowski, P. / Urnavicius, L. / Boland, A. / Fagiewicz, R. / Busselez, J. / Papai, G. / Schmidt, H.|
|Citation||Journal: Elife / Year: 2018|
Title: The CryoEM structure of the ribosome maturation factor Rea1.
Authors: Piotr Sosnowski / Linas Urnavicius / Andreas Boland / Robert Fagiewicz / Johan Busselez / Gabor Papai / Helgo Schmidt
Abstract: The biogenesis of 60S ribosomal subunits is initiated in the nucleus where rRNAs and proteins form pre-60S particles. These pre-60S particles mature by transiently interacting with various assembly ...The biogenesis of 60S ribosomal subunits is initiated in the nucleus where rRNAs and proteins form pre-60S particles. These pre-60S particles mature by transiently interacting with various assembly factors. The ~5000 amino-acid AAA+ ATPase Rea1 (or Midasin) generates force to mechanically remove assembly factors from pre-60S particles, which promotes their export to the cytosol. Here we present three Rea1 cryoEM structures. We visualise the Rea1 engine, a hexameric ring of AAA+ domains, and identify an α-helical bundle of AAA2 as a major ATPase activity regulator. The α-helical bundle interferes with nucleotide-induced conformational changes that create a docking site for the substrate binding MIDAS domain on the AAA +ring. Furthermore, we reveal the architecture of the Rea1 linker, which is involved in force generation and extends from the AAA+ ring. The data presented here provide insights into the mechanism of one of the most complex ribosome maturation factors.
SummaryFull reportAbout validation report
|Date||Deposition: Oct 19, 2018 / Release: Dec 12, 2018|
|Structure viewer||Molecule: |
Downloads & links
|#1: Protein/peptide|| |
Mass: 192045.828 Da / Num. of mol.: 1
Source: (gene. exp.) Saccharomyces cerevisiae (baker's yeast)
Strain: ATCC 204508 / S288c / Gene: MDN1, REA1, YLR106C, L2901, L8004.13 / Production host: Saccharomyces cerevisiae (baker's yeast) / References: UniProt: Q12019
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / Reconstruction method: single particle reconstruction|
|Component||Name: Rea1 (MIDASIN) tail with ADP / Type: COMPLEX / Entity ID: 1 / Source: RECOMBINANT|
|Source (natural)||Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)|
|Source (recombinant)||Organism: Saccharomyces cerevisiae (baker's yeast)|
|Buffer solution||Details: ADP was added 5 minute before the plunging / pH: 7.2|
|Specimen||Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Grid material: COPPER / Grid mesh size: 200 / Grid type: Quantifoil R2/2|
|Vitrification||Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Microscopy||Microscope model: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 105000 / Calibrated magnification: 105000 / Nominal defocus max: 3400 nm / Nominal defocus min: 1800 nm / Cs: 0.01 mm / Alignment procedure: ZEMLIN TABLEAU|
|Specimen holder||Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Residual tilt: 12 mradians|
|Image recording||Average exposure time: 0.2 sec. / Electron dose: 46.2 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number of grids imaged: 14 / Number of real images: 23230|
|EM imaging optics||Energyfilter name: GIF Quantum LS / Energyfilter slit width: 20 eV / Sph aberration corrector: Titan Krios Cs Corrector|
|Image scans||Sampling size: 5 microns / Width: 3712 / Height: 3840 / Movie frames/image: 35 / Used frames/image: 2-35|
|Software||Name: PHENIX / Version: 1.13_2998: / Classification: refinement|
|CTF correction||Type: PHASE FLIPPING AND AMPLITUDE CORRECTION|
|Symmetry||Point symmetry: C1|
|3D reconstruction||Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 432556 / Symmetry type: POINT|
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