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- EMDB-9759: Cryo-EM density map of methionine aminopeptidase enzyme and chape... -

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Basic information

Entry
Database: EMDB / ID: EMD-9759
TitleCryo-EM density map of methionine aminopeptidase enzyme and chaperone trigger factor bound to the E. coli 70S ribosome
Map dataCryo-EM structure of E. coli 70S ribosome in complex with enzyme methionine aminopeptidase and chaperone trigger factor
Sample
  • Complex: E. coli 70S ribosome in complex with methionine aminopeptidase enzyme and chaperone trigger factor
    • Protein or peptide: Methionine aminopeptidaseMethionyl aminopeptidase
    • Protein or peptide: Trigger factor
KeywordsE. coli 70S ribosome / Protein biogenesis / Chaperone / Methionine aminopeptidase / Trigger factor / Polypeptide exit tunnel / RIBOSOME
Function / homology
Function and homology information


'de novo' cotranslational protein folding / initiator methionyl aminopeptidase activity / methionyl aminopeptidase / metalloaminopeptidase activity / protein unfolding / chaperone-mediated protein folding / protein folding chaperone / regulation of DNA-templated transcription elongation / peptidylprolyl isomerase / peptidyl-prolyl cis-trans isomerase activity ...'de novo' cotranslational protein folding / initiator methionyl aminopeptidase activity / methionyl aminopeptidase / metalloaminopeptidase activity / protein unfolding / chaperone-mediated protein folding / protein folding chaperone / regulation of DNA-templated transcription elongation / peptidylprolyl isomerase / peptidyl-prolyl cis-trans isomerase activity / ferrous iron binding / ribosome binding / protein transport / cell cycle / cell division / proteolysis / DNA binding / cytosol / cytoplasm
Similarity search - Function
Transcription elongation factor GreA/GreB, C-terminal domain superfamily / Trigger factor / Trigger factor, C-terminal / Trigger factor, ribosome-binding, bacterial / Trigger factor ribosome-binding domain superfamily / Bacterial trigger factor protein (TF) / Bacterial trigger factor protein (TF) C-terminus / Trigger factor, C-terminal domain superfamily / Trigger factor/SurA domain superfamily / Methionine aminopeptidase subfamily 1 signature. ...Transcription elongation factor GreA/GreB, C-terminal domain superfamily / Trigger factor / Trigger factor, C-terminal / Trigger factor, ribosome-binding, bacterial / Trigger factor ribosome-binding domain superfamily / Bacterial trigger factor protein (TF) / Bacterial trigger factor protein (TF) C-terminus / Trigger factor, C-terminal domain superfamily / Trigger factor/SurA domain superfamily / Methionine aminopeptidase subfamily 1 signature. / Peptidase M24A, methionine aminopeptidase, subfamily 1 / Peptidase M24, methionine aminopeptidase / Peptidase M24 / Metallopeptidase family M24 / Creatinase/aminopeptidase-like
Similarity search - Domain/homology
Methionine aminopeptidase / Trigger factor
Similarity search - Component
Biological speciesEscherichia coli K-12 (bacteria) / Thermotoga maritima MSB8 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 14.2 Å
AuthorsSengupta J / Bhakta S
Funding support India, 2 items
OrganizationGrant numberCountry
Council of Scientific & Industrial Research India
Department of Science & Technology (India)SB/SO/BB-0025/2014 India
CitationJournal: J Mol Biol / Year: 2019
Title: Cryo-EM Structures Reveal Relocalization of MetAP in the Presence of Other Protein Biogenesis Factors at the Ribosomal Tunnel Exit.
Authors: Sayan Bhakta / Shirin Akbar / Jayati Sengupta /
Abstract: During protein biosynthesis in bacteria, one of the earliest events that a nascent polypeptide chain goes through is the co-translational enzymatic processing. The event includes two enzymatic ...During protein biosynthesis in bacteria, one of the earliest events that a nascent polypeptide chain goes through is the co-translational enzymatic processing. The event includes two enzymatic pathways: deformylation of the N-terminal methionine by the enzyme peptide deformylase (PDF), followed by methionine excision catalyzed by methionine aminopeptidase (MetAP). During the enzymatic processing, the emerging nascent protein likely remains shielded by the ribosome-associated chaperone trigger factor. The ribosome tunnel exit serves as a stage for recruiting proteins involved in maturation processes of the nascent chain. Co-translational processing of nascent chains is a critical step for subsequent folding and functioning of mature proteins. Here, we present cryo-electron microscopy structures of Escherichia coli (E. coli) ribosome in complex with the nascent chain processing proteins. The structures reveal overlapping binding sites for PDF and MetAP when they bind individually at the tunnel exit site, where L22-L32 protein region provides primary anchoring sites for both proteins. In the absence of PDF, trigger factor can access ribosomal tunnel exit when MetAP occupies its primary binding site. Interestingly, however, in the presence of PDF, when MetAP's primary binding site is already engaged, MetAP has a remarkable ability to occupy an alternative binding site adjacent to PDF. Our study, thus, discloses an unexpected mechanism that MetAP adopts for context-specific ribosome association.
History
DepositionDec 22, 2018-
Header (metadata) releaseApr 17, 2019-
Map releaseApr 17, 2019-
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: 1
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 1
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6j0a
  • Surface level: 1
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6j0a
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_9759.png

Downloads & links

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Map

FileDownload / File: emd_9759.map.gz / Format: CCP4 / Size: 40.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM structure of E. coli 70S ribosome in complex with enzyme methionine aminopeptidase and chaperone trigger factor
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.89 Å/pix.
x 220 pix.
= 415.8 Å		1.89 Å/pix.
x 220 pix.
= 415.8 Å		1.89 Å/pix.
x 220 pix.
= 415.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.89 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.0 / Movie #1: 1
Minimum - Maximum-8.475949 - 13.180147
Average (Standard dev.)0.072663575 (±1.2372191)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions220220220
Spacing220220220
CellA=B=C: 415.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.891.891.89
M x/y/z220220220
origin x/y/z0.0000.0000.000
length x/y/z415.800415.800415.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS220220220
D min/max/mean-8.47613.1800.073

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Supplemental data

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Sample components

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Entire : E. coli 70S ribosome in complex with methionine aminopeptidase en...

EntireName: E. coli 70S ribosome in complex with methionine aminopeptidase enzyme and chaperone trigger factor
Components
  • Complex: E. coli 70S ribosome in complex with methionine aminopeptidase enzyme and chaperone trigger factor
    • Protein or peptide: Methionine aminopeptidaseMethionyl aminopeptidase
    • Protein or peptide: Trigger factor

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Supramolecule #1: E. coli 70S ribosome in complex with methionine aminopeptidase en...

SupramoleculeName: E. coli 70S ribosome in complex with methionine aminopeptidase enzyme and chaperone trigger factor
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli K-12 (bacteria) / Strain: K-12

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Macromolecule #1: Methionine aminopeptidase

MacromoleculeName: Methionine aminopeptidase / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: methionyl aminopeptidase
Source (natural)Organism: Escherichia coli K-12 (bacteria) / Strain: K-12
Molecular weightTheoretical: 29.341775 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MAISIKTPED IEKMRVAGRL AAEVLEMIEP YVKPGVSTGE LDRICNDYIV NEQHAVSACL GYHGYPKSVC ISINEVVCHG IPDDAKLLK DGDIVNIDVT VIKDGFHGDT SKMFIVGKPT IMGERLCRIT QESLYLALRM VKPGINLREI GAAIQKFVEA E GFSVVREY ...String:
MAISIKTPED IEKMRVAGRL AAEVLEMIEP YVKPGVSTGE LDRICNDYIV NEQHAVSACL GYHGYPKSVC ISINEVVCHG IPDDAKLLK DGDIVNIDVT VIKDGFHGDT SKMFIVGKPT IMGERLCRIT QESLYLALRM VKPGINLREI GAAIQKFVEA E GFSVVREY CGHGIGQGFH EEPQVLHYDS RETNVVLKPG MTFTIEPMVN AGKKEIRTMK DGWTVKTKDR SLSAQYEHTI VV TDNGCEI LTLRKDDTIP AIISHDE

UniProtKB: Methionine aminopeptidase

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Macromolecule #2: Trigger factor

MacromoleculeName: Trigger factor / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: peptidylprolyl isomerase
Source (natural)Organism: Thermotoga maritima MSB8 (bacteria) / Strain: MSB8
Molecular weightTheoretical: 47.961543 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MEVKELERDK NRVVLEYVFG AEEIAQAEDK AVRYLNQRVE IPGFRKGRIP KNVLKMKLGE EFQEYTLDFL MDLIPDTLKD RKLILSPIV TERELKDVTA RVVVEVHEEP EVRIGDISKI EVEKVDEEKV LEKYVERRIE DLRESHALLE PKEGPAEAGD L VRVNMEVY ...String:
MEVKELERDK NRVVLEYVFG AEEIAQAEDK AVRYLNQRVE IPGFRKGRIP KNVLKMKLGE EFQEYTLDFL MDLIPDTLKD RKLILSPIV TERELKDVTA RVVVEVHEEP EVRIGDISKI EVEKVDEEKV LEKYVERRIE DLRESHALLE PKEGPAEAGD L VRVNMEVY NEEGKKLTSR EYEYVISEDE DRPFVKDLVG KKKGDVVEIE REYEGKKYTY KLEVEEVYKR TLPEIGDELA KS VNNEFET LEQLKESLKK EGKEIYDVEM KESMREQLLE KLPEIVEIEI SDRTLEILVN EAINRLKREG RYEQIVSSYE SEE KFREEL KERILDDIKR DRVIEVLAQE KGISVNDEEL EKEAEELAPF WGISPDRAKS LVKARQDLRE ELRWAILKRK VLDL LLQEV E

UniProtKB: Trigger factor

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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: 7.4
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI POLARA 300
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Sample stageCooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI EAGLE (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Average electron dose: 10.0 e/Å2
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionResolution.type: BY AUTHOR / Resolution: 14.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: SPIDER / Number images used: 19500
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
PDB IDChain

1c21

chain_id: A, source_name: PDB, initial_model_type: experimental model

3gu0

chain_id: A, source_name: PDB, initial_model_type: experimental model
Output model

PDB-6j0a:
Crystal structure of E. coli methionine aminopeptidase enzyme and chaperone trigger factor fitted into the cryo-EM density map of the complex

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