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New mRNA-tRNA translocation intermediates revealed by cryoEM: Class 4A (partially rotated (I) 70S ribosome with A-site and P-site tRNAs)

by single particle reconstruction, at 14.7 A resolution

Movie

Orientation:

#1: Surface view with section colored by density value, Surface level: 0.15, Made by UCSF CHIMERA

#2: Surface view colored by height, Surface level: 0.15, Made by UCSF CHIMERA

#3: Surface view with fitted model, atomic models: PDB-3j0v, PDB-3j0w, Surface level: 0.15, Made by UCSF CHIMERA

#4: Simplified surface model with fitted atomic model: PDB-3j0v, Made by Jmol

#5: Simplified surface model with fitted atomic model: PDB-3j0w, Made by Jmol

Entry
Summary
Database / IDEM DATA BANK (EMDB) / 5359
TitleNew mRNA-tRNA translocation intermediates revealed by cryoEM: Class 4A (partially rotated (I) 70S ribosome with A-site and P-site tRNAs)
MapCryo-revealed 70S ribosome partially rotated (i) (Class 4A)
SampleTrp-tRNA-EFTu-GDP-kir-70S ribosome
Keywordsribosome, 70S, partially rotated, intermediates, translocation, tRNA
AuthorsAgirrezabala X, Liao HY, Schreiner E, Fu J, Ortiz-Meoz RF, Schulten K, Green R, Frank J
DateDeposition: 2011-12-02, Header release: 2012-01-26, Map release: 2012-02-16, Last update: 2013-03-06
EMDB SitesEMDB @PDBe (EU), EMDB @RCSB (USA)
Structure Visualization
MoviesMovie Page

#1: Surface view with section colored by density value, Surface level: 0.15, Made by UCSF CHIMERA

#2: Surface view colored by height, Surface level: 0.15, Made by UCSF CHIMERA

#3: Surface view with fitted model, atomic models: PDB-3j0v, PDB-3j0w, Surface level: 0.15, Made by UCSF CHIMERA

#4: Simplified surface model with fitted atomic model: PDB-3j0v, Made by Jmol

#5: Simplified surface model with fitted atomic model: PDB-3j0w, Made by Jmol

Supplemental images

EMDB figure
Structure viewersYorodumi, Launch PeppeR (About PeppeR), Volume viewer (RCSB, PDBe)
Related Structure Data
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Cite: data citing same article

Fit: output model of fitting

Similar strucutres (beta)

Diagram

EMDB-5361

EMDB-1541

EMDB-1363

EMDB-1056

EMDB-1565

EMDB-1006
List of similar structure data about Omokage system
Article
Citation - Primary
ArticleProc. Natl. Acad. Sci. U.S.A., Vol. 109, Issue 16, Page 6094-9, Year 2012
TitleStructural characterization of mRNA-tRNA translocation intermediates.
AuthorsXabier Agirrezabala, Hstau Y Liao, Eduard Schreiner, Jie Fu, Rodrigo F Ortiz-Meoz, Klaus Schulten, Rachel Green, Joachim Frank
Structural Biology Unit, CIC-bioGUNE, Bizkaia Technology Park, Derio 48160, Basque Country, Spain.
KeywordsBinding Sites, Cryoelectron Microscopy, Crystallography, X-Ray, Escherichia coli (genetics), Macromolecular Substances (chemistry), Models, Molecular, Molecular Conformation, Protein Biosynthesis, RNA, Messenger (genetics), RNA, Transfer (genetics, 9014-25-9), Ribosomes (metabolism), Thermodynamics
LinksDOI: 10.1073/pnas.1201288109, PubMed: 22467828, PMC: PMC3340995
Map
FileEMD-5359.map ( map file in CCP4 format, 62502 KB )
Projections & SlicesSize of images:
AxesZ (Sec.)Y (Row.)X (Col.)
Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Density

Histogram

Histogram (log scale)
Contour Level:0.1 (by author), 0.15 (movie #1):
Minimum - Maximum: -0.45961869 - 1.42249143
Average (Standard dev.): 0.02930420 (0.19532192)
Data TypeImage stored as Reals
Space Group Number1
Map Geometry
Axis Order : X Y Z
Dimensions : 250 250 250
Origin : -124 -124 -124
Limit : 125 125 125
Spacing : 250 250 250
Unit CellA = 375.0 A , B = 375.0 A , C = 375.0 A ,
alpha = 90.0 degrees , beta = 90.0 degrees , gamma = 90.0 degrees
Pixel SpacingX = 1.5 A , Y = 1.5 A , Z = 1.5 A
CCP4 map header info
modeImage stored as Reals
A/pix X/Y/Z1.51.51.5
M x/y/z250250250
origin x/y/z0.0000.0000.000
length x/y/z375.000375.000375.000
alpha/beta/gamma90.00090.00090.000
start NX/NY/NZ-62-62-62
NX/NY/NZ125125125
MAP C/R/S123
start NC/NR/NS-124-124-124
NC/NR/NS250250250
start NC,NX/NR,NY/NS,NZ
NC,NX/NR,NY/NS,NZ
D min/max/mean-0.4601.4220.029
Annotation DetailsCryo-revealed 70S ribosome partially rotated (i) (Class 4A)
Supplement
Sample
NameTrp-tRNA-EFTu-GDP-kir-70S ribosome
Number of Components3
Oligomeric Statemonomeric
Theoretical Mass2.8 MDa
Experimental Mass2.8 MDa
Component #1: ribosome-prokaryote - 70S ribosome
Scientific name70S ribosome
Scientific Name of SpeciesEscherichia coli (NCBI Taxonomy: 562)

ProkaryoteALL
Recombinant expressionNo
Experiment
Sample Preparation
Specimen Stateparticle
Specimen Conc0.03 mg/ml
Specimen Support Details200 mesh
BufferDetails: HiFi buffer (50 mM Tris-HCl, pH 7.5, 70mM NH4Cl, 30 mM KCl, 3.5 mM MgCl2, 0.5 mM spermidine, 8mM putrescine, 2 mM DTT, 3.5 mM MgCl2)
pH: 7.5
Vitrification
Methodblot for 3 seconds
Cryogen NameNITROGEN
DetailsVitrification instrument: Vitrobot
Humidity90
InstrumentFEI VITROBOT
Temperature80 Kelvin
Imaging
MicroscopeFEI TECNAI F30
Date01-MAY-2007
Detailsautomated data collection system AutoEMation (CCD mag. 100000x)
Electron Gun
Electron SourceFIELD EMISSION GUN
Accelerating Voltage300 kV
Electron Dose25 e/A**2
Electron Beam Tilt Paramsno tilt
Illumination ModeFLOOD BEAM
Lens
MagnificationNominal: 59000 X, Calibrated: 58269 X
Astigmatismobjective corrected at 100,000 times magnification
Nominal Cs2.26 mm
Imaging ModeBRIGHT FIELD
Defocus1200 nm - 4000 nm
Energy FilterType: no filter , Window:
Specimen Holder
Holdercartridge ( OTHER )
Tilt Angle0 degrees - 0 degrees
Temperature80.7 Kelvin ( 80.7 Kelvin - 80.7 Kelvin )
Camera
DetectorTVIPS TemCam-F415 (CCD)
Image Acquisition
Processing
Methodsingle particle reconstruction
3 D reconstruction
SoftwareXMIPP, SPIDER
CTF Correctiondefocus groups, Wiener filter
DetailsClassification using ML3D
Resolution By Author14.7
Resolution MethodFSC at 0.5 cut-off
Single Particle
Applied SymmetryC1 (asymmetric)
Number of Projections21000
Atomic Model Fitting
Model #0
DetailsProtocol: Molecular Dynamics based flexible fitting
Refinement Protocolflexible
Refinement SpaceREAL
Target CriteriaRMSD, cross correlation
SoftwareMDFF
Model #1
Refinement SpaceREAL
Target CriteriaRMSD, cross correlation
Refinement Protocolflexible
DetailsProtocol: Molecular Dynamics based flexible fitting
SoftwareMDFF
Fitted Coordinate
PDB entry ID

PDB-3j0v

PDB-3j0w
Download
Data from EMDB
Header (meta data in XML format)emd-5359.xml (8.4 KB)
Map dataemd_5359.map.gz (54.8 MB)
FTP directoryftp://ftp.pdbj.org/pub/emdb/structures/EMD-5359
Movie files
movie #1
.mp4 (H.264/MPEG-4 AVC format), 3.3 MB
.webm (WebM/VP8 format), 5.3 MB
Session file for UCSF-Chimera, 26.8 KB
movie #2
.mp4 (H.264/MPEG-4 AVC format), 3 MB
.webm (WebM/VP8 format), 4.4 MB
Session file for UCSF-Chimera, 26.9 KB
movie #3
.mp4 (H.264/MPEG-4 AVC format), 3.8 MB
.webm (WebM/VP8 format), 5.4 MB
Session file for UCSF-Chimera, 41.4 MB
movie #4
.mp4 (H.264/MPEG-4 AVC format), 3.5 MB
.webm (WebM/VP8 format), 4.4 MB
movie #5
.mp4 (H.264/MPEG-4 AVC format), 3.5 MB
.webm (WebM/VP8 format), 4.5 MB