EMDB-1717: Ribosome dynamics and tRNA movement as visualized by t...

Entry
Summary
Database / ID	EM DATA BANK (EMDB) / 1717
Authors	Fischer N, Konevega AL, Wintermeyer W, Rodnina MV, Stark H
EMDB Sites	EMDB @PDBe (EU), EMDB @RCSB (USA)
Structure Visualization
Movies	Movie Page #1: Surface view with section colored by density value, Surface level: 22, Made by UCSF CHIMERA #2: Surface view colored by height, Surface level: 22, Made by UCSF CHIMERA
Supplemental images	image1717.png
Structure viewers	Yorodumi, Launch PeppeR (About PeppeR), Volume viewer (RCSB, PDBe)
Related Structure Data
Related Entries	EMDB-1716 Cite EMDB-1718 Cite EMDB-1719 Cite EMDB-1720 Cite EMDB-1721 Cite EMDB-1722 Cite EMDB-1723 Cite EMDB-1724 Cite EMDB-1725 Cite EMDB-1726 Cite EMDB-1727 Cite Cite: data citing same article
Similar strucutres (beta)	Diagram EMDB-1524 EMDB-1716 EMDB-1721 EMDB-1720 EMDB-1261 EMDB-1564 List of similar structure data about Omokage system

Article
Citation - Primary
Article	Nature, Vol. 466, Issue 7304, Page 329-33, Year 2010
Title	Ribosome dynamics and tRNA movement by time-resolved electron cryomicroscopy.
Authors	Niels Fischer, Andrey L Konevega, Wolfgang Wintermeyer, Marina V Rodnina, Holger Stark 3D Electron Cryomicroscopy Group, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany.
Keywords	Cryoelectron Microscopy, Escherichia coli, Kinetics, Models, Molecular, Molecular Conformation, Movement, Protein Biosynthesis, RNA, Transfer (genetics, 9014-25-9), Ribosome Subunits, Large, Bacterial (chemistry), Ribosome Subunits, Small, Bacterial (chemistry), Ribosomes (chemistry), Temperature, Thermodynamics, Time Factors
Links	DOI: 10.1038/nature09206, PubMed: 20631791

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 = 358.4 A , B = 358.4 A , C = 358.4 A ,
alpha = 90 degrees , beta = 90 degrees , gamma = 90 degrees

Pixel Spacing

X = 2.8 A , Y = 2.8 A , Z = 2.8 A

CCP4 map header info

Annotation Details

Substate of E. coli 70S-fMetVal-tRNAVal-tRNAfMet complex in intermediate pre-translocation state (pre2)

Sample
Name	Substate of E. coli 70S-fMetVal-tRNAVal-tRNAfMet complex in intermediate pre-translocation state (pre2)
Number of Components	4
Theoretical Mass	2.5 MDa
Component #1: ribosome-prokaryote - E. coli 70S
Scientific name	Ribosome
Common Name	E. coli 70S
Theoretical Mass	2.5 MDa
Scientific Name of Species	Escherichia coli (NCBI Taxonomy: 562)
Prokaryote	ALL
Recombinant expression	No
Component #2: nucleic-acid - peptidyl tRNA
Scientific name	fMetVal-tRNAVal
Common Name	peptidyl tRNA
Theoretical Mass	0.025 MDa
Scientific Name of Species	Escherichia coli (NCBI Taxonomy: 562)
Class	T-RNA
Structure	DOUBLE HELIX
Synthetic	No
Component #3: nucleic-acid - deacylated tRNA
Scientific name	tRNAfMet
Common Name	deacylated tRNA
Theoretical Mass	0.025 MDa
Scientific Name of Species	Escherichia coli (NCBI Taxonomy: 562)
Class	T-RNA
Structure	DOUBLE HELIX
Synthetic	No
Component #4: nucleic-acid - mRNA
Scientific name	m022 mRNA
Common Name	mRNA
Details	Coding sequence AUGGUU
Scientific Name of Species	synthetic construct (NCBI Taxonomy: 32630)
Structure	SINGLE STRANDED
Synthetic	Yes
Class	RNA

Experiment
Sample Preparation
Specimen State	particle
Buffer	Details: 50 mM Tris-HCl, 70 mM NH4Cl, 30 mM KCl, 7 mM MgCl2, 0.6 mM spermine, 0.4 mM spermidine pH: 7.5
Vitrification
Method	Manual blotting for about 2 seconds
Cryogen Name	ETHANE
Time Resolved State	Samples were vitrified at different time points along the reaction coordinate (1, 2, 5 and 20 minutes after addition of deacylated tRNAfMet to 70S-fMetVal-tRNAVal complexes)
Details	Vitrification instrument: Custom-built CEVS. Dew-point temperature (temperature on the grid) adjusted to 18 degrees C
Humidity	75
Instrument	HOMEMADE PLUNGER
Temperature	77 Kelvin
Imaging
Microscope	FEI/PHILIPS CM200FEG
Electron Gun
Electron Source	FIELD EMISSION GUN
Accelerating Voltage	160 kV
Electron Dose	20 e/A**2
Illumination Mode	SPOT SCAN
Lens
Magnification	Nominal: 161000 X, Calibrated: 162740 X
Astigmatism	Objective lens astigmatism was corrected at 200,000 times magnification
Nominal Cs	2.0 mm
Imaging Mode	BRIGHT FIELD
Defocus	500 nm - 2000 nm
Specimen Holder
Holder	Eucentric ( GATAN LIQUID NITROGEN )
Temperature	77 Kelvin
Camera
Detector	4k CCD camera (TVIPS)
Image Acquisition

Processing
Method	single particle reconstruction
3 D reconstruction
Algorithm	Projection matching
Software	IMAGIC, custom, Spider
CTF Correction	local
Details	Final maps were calculated from 13 datasets acquired at different time points, computationally sorted into distinct substates
Resolution By Author	20
Resolution Method	FSC at 0.5 cut-off
Single Particle
Number of Projections	1904

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), 3.4 MB .webm (WebM/VP8 format), 5.4 MB Session file for UCSF-Chimera, 26.5 KB

movie #2

	.mp4 (H.264/MPEG-4 AVC format), 3.2 MB .webm (WebM/VP8 format), 4.7 MB Session file for UCSF-Chimera, 26.7 KB