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- EMDB-47518: Escherichia coli encapsulin-associated DyP peroxidase -

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Basic information

Entry
Database: EMDB / ID: EMD-47518
TitleEscherichia coli encapsulin-associated DyP peroxidase
Map data
Sample
  • Complex: Escherichia coli KTE40 encapsulin-associated DyP peroxidase
    • Protein or peptide: Dyp-type peroxidase
  • Ligand: PROTOPORPHYRIN IX CONTAINING FE
KeywordsDye-decolorizing peroxidase / peroxidase / DyP / encapsulin / OXIDOREDUCTASE
Function / homology
Function and homology information


peroxidase activity / heme binding / cytosol
Similarity search - Function
: / : / Dyp-type peroxidase, C-terminal / Dyp-type peroxidase, N-terminal / DyP-type peroxidase family. / Dyp-type peroxidase / Dimeric alpha-beta barrel
Similarity search - Domain/homology
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.31 Å
AuthorsAndreas MP / Ubilla NC / Giessen TW
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS) United States
CitationJournal: bioRxiv / Year: 2024
Title: Structural and biochemical characterization of a widespread enterobacterial peroxidase encapsulin.
Authors: Natalia C Ubilla-Rodriguez / Michael P Andreas / Tobias W Giessen /
Abstract: Encapsulins are self-assembling protein compartments found in prokaryotes and specifically encapsulate dedicated cargo enzymes. The most abundant encapsulin cargo class are Dye-decolorizing ...Encapsulins are self-assembling protein compartments found in prokaryotes and specifically encapsulate dedicated cargo enzymes. The most abundant encapsulin cargo class are Dye-decolorizing Peroxidases (DyPs). It has been previously suggested that DyP encapsulins are involved in oxidative stress resistance and bacterial pathogenicity due to DyPs' inherent ability to reduce and detoxify hydrogen peroxide while oxidizing a broad range of organic co-substrates. Here, we report the structural and biochemical analysis of a DyP encapsulin widely found across enterobacteria. Using bioinformatic approaches, we show that this DyP encapsulin is encoded by a conserved transposon-associated operon, enriched in enterobacterial pathogens. Through low pH and peroxide exposure experiments, we highlight the stability of this DyP encapsulin under harsh conditions and show that DyP catalytic activity is highest at low pH. We determine the structure of the DyP-loaded shell and free DyP via cryo-electron microscopy, revealing the structural basis for DyP cargo loading and peroxide preference. Our work lays the foundation to further explore the substrate range and physiological functions of enterobacterial DyP encapsulins.
History
DepositionOct 25, 2024-
Header (metadata) releaseMar 19, 2025-
Map releaseMar 19, 2025-
UpdateMar 19, 2025-
Current statusMar 19, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_47518.png

Downloads & links

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Map

FileDownload / File: emd_47518.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)X (Row.)Y (Col.)
0.91 Å/pix.
x 256 pix.
= 232.96 Å		0.91 Å/pix.
x 256 pix.
= 232.96 Å		0.91 Å/pix.
x 256 pix.
= 232.96 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.91 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.08
Minimum - Maximum-0.30613226 - 0.51517224
Average (Standard dev.)0.0010788586 (±0.020197608)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 232.96 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_47518_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_47518_half_map_2.map
Projections & Slices
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Histogram

Histogram (log scale)

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Sample components

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Entire : Escherichia coli KTE40 encapsulin-associated DyP peroxidase

EntireName: Escherichia coli KTE40 encapsulin-associated DyP peroxidase
Components
  • Complex: Escherichia coli KTE40 encapsulin-associated DyP peroxidase
    • Protein or peptide: Dyp-type peroxidase
  • Ligand: PROTOPORPHYRIN IX CONTAINING FE

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Supramolecule #1: Escherichia coli KTE40 encapsulin-associated DyP peroxidase

SupramoleculeName: Escherichia coli KTE40 encapsulin-associated DyP peroxidase
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Escherichia coli (E. coli) / Strain: KTE40

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Macromolecule #1: Dyp-type peroxidase

MacromoleculeName: Dyp-type peroxidase / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli) / Strain: KTE40
Molecular weightTheoretical: 40.644516 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MHHHHHHGGS ENLYFQSGGS GCPFSQSVSQ PVDERLTRAA IFLVVTINPG KAAEVAVRAH CSILSSLIRG VGFRISDGGL SCVMGVSEG GWERLFGDTK PEYLHVFREI NGVHHAPSTP GDLLYHIRAA RMDLCFELAS RILSDLGNSV SVVDSVQGFR Y FDDRDLLG ...String:
MHHHHHHGGS ENLYFQSGGS GCPFSQSVSQ PVDERLTRAA IFLVVTINPG KAAEVAVRAH CSILSSLIRG VGFRISDGGL SCVMGVSEG GWERLFGDTK PEYLHVFREI NGVHHAPSTP GDLLYHIRAA RMDLCFELAS RILSDLGNSV SVVDSVQGFR Y FDDRDLLG FVDGTENPVA QAAVDATLIG DEDMVFAGGS YVIVQKYLHD LDKWNAIPVE QQEKIIGREK LSDIELRDAD KP SYAHNVL TSIEEDGEDV DILRDNMPFG DPGKGEFGTY FIGYSRKPER IERMLENMFI GNPPGNYDRI LDVSRAITGT LFF VPTTSF LDSIEPQSAP GQQGDDVINT LRSTAIKGDI MPGSLNIGSL KKEV

UniProtKB: Dyp-type peroxidase

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Macromolecule #2: PROTOPORPHYRIN IX CONTAINING FE

MacromoleculeName: PROTOPORPHYRIN IX CONTAINING FE / type: ligand / ID: 2 / Number of copies: 1 / Formula: HEM
Molecular weightTheoretical: 616.487 Da
Chemical component information

ChemComp-HEM:
PROTOPORPHYRIN IX CONTAINING FE

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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

Concentration0.5 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
150.0 mMNaClsodium chloride
20.0 mMC4H11NO3tris(hydroxymethyl)aminomethane

Details: Sample was prepared in 20 mM Tris pH 7.5, 150 mM NaCl.
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
Details: Grids were glow discharged for 60 seconds at 5 mA in vacuum.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV
Details: Grids were frozen with the following parameters: blot force 20, blot time 4 seconds, humidty 100 percent, 22 degrees C..
DetailsSample was frozen on two grids, one at 0.5 mg/mL and the other at 0.25 mg/mL.

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Electron microscopy #1

Microscopy ID1
MicroscopeFEI TALOS ARCTICA
Image recordingImage recording ID: 1 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Number grids imaged: 1 / Number real images: 123 / Average exposure time: 6.0 sec. / Average electron dose: 47.84 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.8 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 45000
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Electron microscopy #1~

Microscopy ID1
MicroscopeTFS GLACIOS
Image recordingImage recording ID: 2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Number grids imaged: 1 / Number real images: 206 / Average exposure time: 4.0 sec. / Average electron dose: 46.24 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.8 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 45000

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Image processing

Image recording ID1
DetailsParticles were picked from micrographs collected on two different grids using TFS Glacios and FEI Talos Arctica cryogenic electron microscopes.
Particle selectionNumber selected: 115178
Startup modelType of model: INSILICO MODEL / In silico model: Ab-Initio reconstruction
Final reconstructionApplied symmetry - Point group: D3 (2x3 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 3.31 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.5.1) / Software - details: Local Refinement
Details: Local refinement with D3 symmetry and force re-do GS-split applied.
Number images used: 10490
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.5.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.5.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

an60


Chain - Source name: AlphaFold / Chain - Initial model type: in silico model
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 105.9 / Target criteria: cross-correlation coefficient
Output model

PDB-9e4r:
Escherichia coli encapsulin-associated DyP peroxidase

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