EMDB-2008: 3D reconstruction of a translating yeast 80S ribosome ...

Entry
Summary
Database / ID	EM DATA BANK (EMDB) / 2008
Title	3D reconstruction of a translating yeast 80S ribosome in complex with Dom34p and Rli1p
Map	This is a 3D cryo-EM reconstruction of a translating yeast 80S ribosome in complex with Dom34p and Rli1p
Sample	Saccharomyces cerevisiae 80S ribosome stalled by a synthetic stem-loop mRNA in complex with Dom34p and Rli1p
Keywords	translation, ribosome recycling, no-go mRNA decay
Authors	Becker T, Franckenberg S, Wickles S, Shoemaker CJ, Anger AM, Armache J-P, Sieber H, Ungewickell C, Berninghausen O, Daberkow I, Karcher A, Thomm M, Hopfner K-P, Green R, Beckmann R
Date	Deposition: 2011-12-09, Header release: 2012-02-15, Map release: 2012-02-17, Last update: 2012-03-01
EMDB Sites	EMDB @PDBe (EU), EMDB @RCSB (USA)
Structure Visualization
Movies	Movie Page #1: Surface view with section colored by density value, Surface level: 0.32, Made by UCSF CHIMERA #2: Surface view colored by height, Surface level: 0.32, Made by UCSF CHIMERA
Supplemental images	EMBD_2008.gif
Structure viewers	Yorodumi, Launch PeppeR (About PeppeR), Volume viewer (RCSB, PDBe)
Related Structure Data
Related Entries	EMDB-2009 Cite EMDB-2010 Cite PDB-3j15 Cite PDB-3j16 Cite Cite: data citing same article
Similar strucutres (beta)	Diagram EMDB-1768 EMDB-1217 EMDB-1666 EMDB-1665 EMDB-1767 EMDB-1780 List of similar structure data about Omokage system

Article
Citation - Primary
Article	Nature, Vol. 482, Issue 7386, Page 501-6, Year 2012
Title	Structural basis of highly conserved ribosome recycling in eukaryotes and archaea.
Authors	Thomas Becker, Sibylle Franckenberg, Stephan Wickles, Christopher J Shoemaker, Andreas M Anger, Jean-Paul Armache, Heidemarie Sieber, Charlotte Ungewickell, Otto Berninghausen, Ingo Daberkow, Annette Karcher, Michael Thomm, Karl-Peter Hopfner, Rachel Green, Roland Beckmann Gene Center and Center for integrated Protein Science Munich, Department of Biochemistry, University of Munich, Feodor-Lynen-Straße 25, 81377 Munich, Germany.
Keywords	ATP-Binding Cassette Transporters (chemistry), Cell Cycle Proteins (chemistry), Cryoelectron Microscopy, Dom34 protein, S cerevisiae (3.1.-), Endoribonucleases (chemistry, 3.1.-), Evolution, Molecular, Iron-Sulfur Proteins (chemistry), Models, Molecular, Movement, Multiprotein Complexes (chemistry), Nuclear Proteins (chemistry), Peptide Termination Factors (chemistry), Protein Binding, Protein Stability, Protein Structure, Tertiary, Pyrococcus furiosus (chemistry), RLI1 protein, S cerevisiae, Ribosomes (chemistry), Saccharomyces cerevisiae (chemistry), Saccharomyces cerevisiae Proteins (chemistry)
Links	DOI: 10.1038/nature10829, PubMed: 22358840

File

Projections & Slices

Size of images:

Images are generated by Spider package.

Density

Data Type

Image stored as Reals

Space Group Number

Map Geometry

Unit Cell

A = 430.56 A , B = 430.56 A , C = 430.56 A ,
alpha = 90.0 degrees , beta = 90.0 degrees , gamma = 90.0 degrees

Pixel Spacing

CCP4 map header info

Annotation Details

This is a 3D cryo-EM reconstruction of a translating yeast 80S ribosome in complex with Dom34p and Rli1p

Sample
Name	Saccharomyces cerevisiae 80S ribosome stalled by a synthetic stem-loop mRNA in complex with Dom34p and Rli1p
Number of Components	3
Oligomeric State	One 80S ribosome binds one Dom34p and one Rli1p
Component #1: ribosome-eukaryote - Baker's yeast 80S ribosome
Scientific name	S. cerevisiae 80S ribosome
Common Name	Baker's yeast 80S ribosome
Scientific Name of Species	Saccharomyces cerevisiae (NCBI Taxonomy: 4932)
Common Name of Species	baker's yeast
Eukaryote	ALL
Recombinant expression	No
Component #2: protein - ABCE1
Scientific name	Rli1p
Common Name	ABCE1
Oligomeric Details	Monomer
Scientific Name of Species	Saccharomyces cerevisiae (NCBI Taxonomy: 4932)
Common Name of Species	baker's yeast
Recombinant expression	No
Component #3: protein - Pelota
Scientific name	Dom34p
Common Name	Pelota
Oligomeric Details	Monomer
Scientific Name of Species	Saccharomyces cerevisiae (NCBI Taxonomy: 4932)
Common Name of Species	Baker's Yeast
Recombinant expression	No

Experiment
Sample Preparation
Specimen Support Details	Quantifoil grids (3/3) with 2 nm carbon on top
Specimen State	particle
Buffer	Details: 20 mM Tris/HCl pH 7.0, 150 mM KOAc, 10 mM Mg(OAc)2, 1.5 mM DTT, 0.005% Nikkol, 0.01 mg/ml Cycloheximide, 0.3% (w/v) Digitonin, 0.5 mM ADPNP pH: 7.0
Vitrification
Method	Blot for 10 seconds before plunging, use 2 layer of filter paper
Cryogen Name	ETHANE
Details	Vitrification instrument: Vitrobot
Humidity	95
Instrument	FEI VITROBOT
Imaging
Microscope	FEI TITAN KRIOS
Details	Final magnification of the object on the CCD image is 128200
Electron Gun
Electron Source	FIELD EMISSION GUN
Accelerating Voltage	300 kV
Electron Dose	25 e/A**2
Illumination Mode	FLOOD BEAM
Lens
Magnification	Nominal: 75000 X,
Nominal Cs	2.7 mm
Imaging Mode	BRIGHT FIELD
Defocus	1400 nm - 4500 nm
Specimen Holder
Holder	autoloader ( OTHER )
Camera
Detector	Eagle 4k CCD camera
Image Acquisition
Number of Digital Images	10000
Sampling Size	15 microns
Quant Bit Number	16

Processing
Method	single particle reconstruction
3 D reconstruction
Algorithm	projection matching
Euler Angles Details	SPIDER
Software	SPIDER
CTF Correction	Wiener Filter
Details	sorting for ribosome conformation and ligand presence was performed
Resolution By Author	7.2
Resolution Method	FSC at 0.5 cut-off
Single Particle
Number of Projections	45700
Details	mammalian Sec61 complex was added to saturate hydrophobic nascent polypeptide chains
Atomic Model Fitting
Model #0
Details	Protocol: rigid body followed by molecular dynamics flexible fitting. rigid body fitting of individual domains followed by molecular dynamics flexible fitting
Refinement Protocol	flexible
Refinement Space	REAL
Software	MDFF
Model #1
Refinement Space	REAL
Refinement Protocol	flexible
Details	Protocol: rigid body followed by molecular dynamics flexible fitting. rigid body followed by molecular dynamics flexible fitting
Software	MDFF

Data from EMDB

Header (meta data in XML format)

Map data

Images

FTP directory

Movie files

movie #1

	.mp4 (H.264/MPEG-4 AVC format), 3.5 MB .webm (WebM/VP8 format), 5.4 MB Session file for UCSF-Chimera, 26.9 KB

movie #2

	.mp4 (H.264/MPEG-4 AVC format), 3.2 MB .webm (WebM/VP8 format), 5 MB Session file for UCSF-Chimera, 26.5 KB