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- EMDB-4082: Structure of bacterial 30S-IF1-IF2-IF3-mRNA-tRNA translation pre-... -

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Basic information

Entry
Database: EMDB / ID: 4082
TitleStructure of bacterial 30S-IF1-IF2-IF3-mRNA-tRNA translation pre-initiation complex(state-II)
Map data
Sample30S-IF1-IF2-IF3-mRNA-tRNA pre-initiation complex (state-II):
SourceThermus thermophilus HB8 (bacteria)
Methodsingle particle reconstruction / cryo EM / 8.3 Å resolution
AuthorsHussain T / Llacer JL / Wimberly BT / Ramakrishnan V
CitationJournal: Cell / Year: 2016
Title: Large-Scale Movements of IF3 and tRNA during Bacterial Translation Initiation.
Authors: Tanweer Hussain / Jose L Llácer / Brian T Wimberly / Jeffrey S Kieft / V Ramakrishnan
Abstract: In bacterial translational initiation, three initiation factors (IFs 1-3) enable the selection of initiator tRNA and the start codon in the P site of the 30S ribosomal subunit. Here, we report 11 ...In bacterial translational initiation, three initiation factors (IFs 1-3) enable the selection of initiator tRNA and the start codon in the P site of the 30S ribosomal subunit. Here, we report 11 single-particle cryo-electron microscopy (cryoEM) reconstructions of the complex of bacterial 30S subunit with initiator tRNA, mRNA, and IFs 1-3, representing different steps along the initiation pathway. IF1 provides key anchoring points for IF2 and IF3, thereby enhancing their activities. IF2 positions a domain in an extended conformation appropriate for capturing the formylmethionyl moiety charged on tRNA. IF3 and tRNA undergo large conformational changes to facilitate the accommodation of the formylmethionyl-tRNA (fMet-tRNA(fMet)) into the P site for start codon recognition.
DateDeposition: Aug 1, 2016 / Header (metadata) release: Aug 10, 2016 / Map release: Oct 5, 2016 / Last update: Jul 12, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

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Map

Fileemd_4082.map.gz (map file in CCP4 format, 70305 KB)
Projections & slices

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AxesZ (Sec.)Y (Row.)X (Col.)
260 pix
1.34 Å/pix.
= 348.4 Å
260 pix
1.34 Å/pix.
= 348.4 Å
260 pix
1.34 Å/pix.
= 348.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.34 Å
Density
Contour Level:0.05 (by author), 0.05 (movie #1):
Minimum - Maximum-0.08887035 - 0.20115758
Average (Standard dev.)0.0011401811 (0.015841644)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions260260260
Origin000
Limit259259259
Spacing260260260
CellA=B=C: 348.4 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.341.341.34
M x/y/z260260260
origin x/y/z0.0000.0000.000
length x/y/z348.400348.400348.400
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS260260260
D min/max/mean-0.0890.2010.001

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Supplemental data

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Sample components

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Entire 30S-IF1-IF2-IF3-mRNA-tRNA pre-initiation complex (state-II)

EntireName: 30S-IF1-IF2-IF3-mRNA-tRNA pre-initiation complex (state-II)
Number of components: 1
MassTheoretical: 912.9 kDa

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Component #1: protein, 30S-IF1-IF2-IF3-mRNA-tRNA pre-initiation complex (state-II)

ProteinName: 30S-IF1-IF2-IF3-mRNA-tRNA pre-initiation complex (state-II)
Recombinant expression: No
MassTheoretical: 912.9 kDa
SourceSpecies: Thermus thermophilus HB8 (bacteria)

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.065 mg/ml / pH: 6.5
VitrificationInstrument: FEI VITROBOT MARK I / Cryogen name: ETHANE / Temperature: 277 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 30 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 59000 X (nominal), 104478 X (calibrated) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 1500 - 3500 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature: K ( 90 - 100 K)
CameraDetector: FEI FALCON II (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 3200

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 8423
3D reconstructionAlgorithm: FOURIER SPACE / Software: RELION / Resolution: 8.3 Å / Resolution method: FSC 0.143 CUT-OFF

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