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- EMDB-6925: M. smegmatis Trans-translation state 70S ribosome -

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Basic information

Entry
Database: EMDB / ID: EMD-6925
TitleM. smegmatis Trans-translation state 70S ribosome
Map dataTrans-translating 70S map of M. smegmatis
Sample
  • Complex: tmRNA and SmpB in trans-translating state of M. smegmatis
    • RNA: tmRNATransfer-messenger RNA
    • RNA: A-tRNAfMet
    • Protein or peptide: SsrA-binding protein
Keywordstrans-translating state / RNA BINDING PROTEIN-RNA complex
Function / homologytrans-translation / SsrA-binding protein / SsrA-binding protein, conserved site / Small protein B / SmpB protein / SsrA-binding protein. / RNA binding / cytoplasm / SsrA-binding protein
Function and homology information
Biological speciesMycobacterium smegmatis str. MC2 155 (bacteria) / Mycobacterium smegmatis (strain ATCC 700084 / mc(2)155) (bacteria) / Escherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 12.5 Å
AuthorsMishra S / Ahmed T
Funding support Singapore, 1 items
OrganizationGrant numberCountry
Ministry of Education (Singapore) Singapore
CitationJournal: Sci Rep / Year: 2018
Title: Structures of Mycobacterium smegmatis 70S ribosomes in complex with HPF, tmRNA, and P-tRNA.
Authors: Satabdi Mishra / Tofayel Ahmed / Anu Tyagi / Jian Shi / Shashi Bhushan /
Abstract: Ribosomes are the dynamic protein synthesis machineries of the cell. They may exist in different functional states in the cell. Therefore, it is essential to have structural information on these ...Ribosomes are the dynamic protein synthesis machineries of the cell. They may exist in different functional states in the cell. Therefore, it is essential to have structural information on these different functional states of ribosomes to understand their mechanism of action. Here, we present single particle cryo-EM reconstructions of the Mycobacterium smegmatis 70S ribosomes in the hibernating state (with HPF), trans-translating state (with tmRNA), and the P/P state (with P-tRNA) resolved to 4.1, 12.5, and 3.4 Å, respectively. A comparison of the P/P state with the hibernating state provides possible functional insights about the Mycobacteria-specific helix H54a rRNA segment. Interestingly, densities for all the four OB domains of bS1 protein is visible in the hibernating 70S ribosome displaying the molecular details of bS1-70S interactions. Our structural data shows a Mycobacteria-specific H54a-bS1 interaction which seems to prevent subunit dissociation and degradation during hibernation without the formation of 100S dimer. This indicates a new role of bS1 protein in 70S protection during hibernation in Mycobacteria in addition to its conserved function during translation initiation.
History
DepositionFeb 28, 2018-
Header (metadata) releaseSep 26, 2018-
Map releaseSep 26, 2018-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5zey
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-5zey
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_6925.png

Downloads & links

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Map

FileDownload / File: emd_6925.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTrans-translating 70S map of M. smegmatis
Voxel sizeX=Y=Z: 1.05 Å
Density
Contour LevelBy AUTHOR: 0.02 / Movie #1: 0.02
Minimum - Maximum-0.041628912 - 0.08586581
Average (Standard dev.)0.0013222978 (±0.0068636946)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 419.99997 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.051.051.05
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z420.000420.000420.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.0420.0860.001

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Supplemental data

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Sample components

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Entire : tmRNA and SmpB in trans-translating state of M. smegmatis

EntireName: tmRNA and SmpB in trans-translating state of M. smegmatis
Components
  • Complex: tmRNA and SmpB in trans-translating state of M. smegmatis
    • RNA: tmRNATransfer-messenger RNA
    • RNA: A-tRNAfMet
    • Protein or peptide: SsrA-binding protein

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Supramolecule #1: tmRNA and SmpB in trans-translating state of M. smegmatis

SupramoleculeName: tmRNA and SmpB in trans-translating state of M. smegmatis
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Mycobacterium smegmatis str. MC2 155 (bacteria) / Location in cell: cytoplasm
Molecular weightTheoretical: 2.6 MDa

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Macromolecule #1: tmRNA

MacromoleculeName: tmRNA / type: rna / ID: 1 / Number of copies: 1
Source (natural)Organism: Mycobacterium smegmatis (strain ATCC 700084 / mc(2)155) (bacteria)
Strain: ATCC 700084 / mc(2)155
Molecular weightTheoretical: 119.208492 KDa
SequenceString: GGGGCUGAAC GGUUUCGACU UCGAGCAUCG AAUCCAGGGA AGCGUGCCGG UGCAGGCAAG AGACCACCGU AAGCGUCGUU GCAACCAAU UAAGCGCCGA UUCCAAUCAG CGCGACUACG CCCUCGCUGC CUAAGCGACG GCUGGUCUGU CAGACCGGGA G UGCCCUCG ...String:
GGGGCUGAAC GGUUUCGACU UCGAGCAUCG AAUCCAGGGA AGCGUGCCGG UGCAGGCAAG AGACCACCGU AAGCGUCGUU GCAACCAAU UAAGCGCCGA UUCCAAUCAG CGCGACUACG CCCUCGCUGC CUAAGCGACG GCUGGUCUGU CAGACCGGGA G UGCCCUCG GCCCGGAUCC UGGCAUCAGC UAGAGGGACC CACCCACGGG UUCGGUCGCG GGACCUGUGG GGACAUCAAA CA GCGACUG GGAUCGUCAU CUCGGCUUGU UCGUGUGACC GGGAGAUCCG AGUAGAGACA UAGCGAACUG CGCACGGAGA AGC CUCGAG GACAUGCCGU AGGACCCGGG UUCAAUUCCC GGCAGCUCCA CCA

GENBANK: GENBANK: CP000480.1

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Macromolecule #2: A-tRNAfMet

MacromoleculeName: A-tRNAfMet / type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 24.786785 KDa
SequenceString:
CGCGGGGUGG AGCAGCCUGG UAGCUCGUCG GGCUCAUAAC CCGAAGAUCG UCGGUUCAAA UCCGGCCCCC GCAACCA

GENBANK: GENBANK: CP016018.1

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Macromolecule #3: SsrA-binding protein

MacromoleculeName: SsrA-binding protein / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium smegmatis (strain ATCC 700084 / mc(2)155) (bacteria)
Strain: ATCC 700084 / mc(2)155
Molecular weightTheoretical: 18.325039 KDa
SequenceString:
MTKKSASSNN KVVATNRKAR HNYTILDTYE AGIVLMGTEV KSLREGQASL ADAFATVDDG EIWLRNVHIA EYHHGTWTNH APRRNRKLL LHRKQIDNLI GKIRDGNLTL VPLSIYFTDG KVKVELALAR GKQAHDKRQD LARRDAQREV IRELGRRAKG K I

UniProtKB: SsrA-binding protein

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5
GridMaterial: COPPER
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Average electron dose: 1.5 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 12.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 391837

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