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- EMDB-12859: Model of closed pentamer of the Haliangium ochraceum encapsulin f... -

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Basic information

Entry
Database: EMDB / ID: EMD-12859
TitleModel of closed pentamer of the Haliangium ochraceum encapsulin from symmetry expansion of icosahedral single particle reconstruction
Map dataRefine 3D map
Sample
  • Complex: Ternary complex of Haliangium ochraceum encapsulin and encapsulated ferritin proteins
    • Protein or peptide: Linocin_M18 bacteriocin protein
    • Protein or peptide: Haliangium ochraceum Encapsulated ferritin localisation sequence
  • Ligand: water
Function / homology
Function and homology information


encapsulin nanocompartment / ferroxidase / ferroxidase activity / iron ion transport / intracellular iron ion homeostasis / metal ion binding
Similarity search - Function
Ferritin-like protein / Type 1 encapsulin shell protein / Encapsulating protein for peroxidase / Ferritin-like superfamily
Similarity search - Domain/homology
Encapsulated ferritin-like protein / Type 1 encapsulin shell protein
Similarity search - Component
Biological speciesHaliangium ochraceum (bacteria) / Haliangium ochraceum (strain DSM 14365 / JCM 11303 / SMP-2) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.4 Å
AuthorsMarles-Wright J / Basle A / Clarke DJ / Ross J
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC)BB/N005570/1 United Kingdom
Citation
Journal: Sci Adv / Year: 2022
Title: Pore dynamics and asymmetric cargo loading in an encapsulin nanocompartment.
Authors: Jennifer Ross / Zak McIver / Thomas Lambert / Cecilia Piergentili / Jasmine Emma Bird / Kelly J Gallagher / Faye L Cruickshank / Patrick James / Efrain Zarazúa-Arvizu / Louise E Horsfall / ...Authors: Jennifer Ross / Zak McIver / Thomas Lambert / Cecilia Piergentili / Jasmine Emma Bird / Kelly J Gallagher / Faye L Cruickshank / Patrick James / Efrain Zarazúa-Arvizu / Louise E Horsfall / Kevin J Waldron / Marcus D Wilson / C Logan Mackay / Arnaud Baslé / David J Clarke / Jon Marles-Wright /
Abstract: Encapsulins are protein nanocompartments that house various cargo enzymes, including a family of decameric ferritin-like proteins. Here, we study a recombinant encapsulin:encapsulated ferritin ...Encapsulins are protein nanocompartments that house various cargo enzymes, including a family of decameric ferritin-like proteins. Here, we study a recombinant encapsulin:encapsulated ferritin complex using cryo-electron microscopy and hydrogen/deuterium exchange mass spectrometry to gain insight into the structural relationship between the encapsulin shell and its protein cargo. An asymmetric single-particle reconstruction reveals four encapsulated ferritin decamers in a tetrahedral arrangement within the encapsulin nanocompartment. This leads to a symmetry mismatch between the protein cargo and the icosahedral encapsulin shell. The encapsulated ferritin decamers are offset from the interior face of the encapsulin shell. Using hydrogen/deuterium exchange mass spectrometry, we observed the dynamic behavior of the major fivefold pore in the encapsulin shell and show the pore opening via the movement of the encapsulin A-domain. These data will accelerate efforts to engineer the encapsulation of heterologous cargo proteins and to alter the permeability of the encapsulin shell via pore modifications.
#1: Journal: Biorxiv / Year: 2021
Title: Model of Haliangium ochraceum encapsulin from icosahedral single particle reconstruction
Authors: Marles-Wright J / Basle A / Clarke DJ / Ross J
History
DepositionMay 1, 2021-
Header (metadata) releaseFeb 9, 2022-
Map releaseFeb 9, 2022-
UpdateFeb 9, 2022-
Current statusFeb 9, 2022Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7oe2
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7oe2
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_12859.png

Downloads & links

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Map

FileDownload / File: emd_12859.map.gz / Format: CCP4 / Size: 729 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationRefine 3D map
Voxel sizeX=Y=Z: 0.652 Å
Density
Contour LevelBy AUTHOR: 0.01 / Movie #1: 0.01
Minimum - Maximum-0.010531169 - 0.028896166
Average (Standard dev.)-0.00039376642 (±0.0014212929)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions576576576
Spacing576576576
CellA=B=C: 375.552 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.6520.6520.652
M x/y/z576576576
origin x/y/z0.0000.0000.000
length x/y/z375.552375.552375.552
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS576576576
D min/max/mean-0.0110.029-0.000

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Supplemental data

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Mask #1

Fileemd_12859_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Post processed masked map

Fileemd_12859_additional_1.map
AnnotationPost processed masked map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half 1

Fileemd_12859_half_map_1.map
AnnotationHalf 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half 2

Fileemd_12859_half_map_2.map
AnnotationHalf 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Ternary complex of Haliangium ochraceum encapsulin and encapsulat...

EntireName: Ternary complex of Haliangium ochraceum encapsulin and encapsulated ferritin proteins
Components
  • Complex: Ternary complex of Haliangium ochraceum encapsulin and encapsulated ferritin proteins
    • Protein or peptide: Linocin_M18 bacteriocin protein
    • Protein or peptide: Haliangium ochraceum Encapsulated ferritin localisation sequence
  • Ligand: water

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Supramolecule #1: Ternary complex of Haliangium ochraceum encapsulin and encapsulat...

SupramoleculeName: Ternary complex of Haliangium ochraceum encapsulin and encapsulated ferritin proteins
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2 / Details: Complex produced by co-expression in E. coli
Source (natural)Organism: Haliangium ochraceum (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
Molecular weightTheoretical: 1.8 MDa

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Macromolecule #1: Linocin_M18 bacteriocin protein

MacromoleculeName: Linocin_M18 bacteriocin protein / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Haliangium ochraceum (strain DSM 14365 / JCM 11303 / SMP-2) (bacteria)
Strain: DSM 14365 / JCM 11303 / SMP-2
Molecular weightTheoretical: 28.844598 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MDLLKRHLAP IVPDAWSAID EEAKEIFQGH LAGRKLVDFR GPFGWEYAAV NTGELRPIDD TPEDVDMKLR QVQPLAEVRV PFTLDVTEL DSVARGATNP DLDDVARAAE RMVEAEDSAI FHGWAQAGIK GIVDSTPHEA LAVASVSDFP RAVLSAADTL R KAGVTGPY ...String:
MDLLKRHLAP IVPDAWSAID EEAKEIFQGH LAGRKLVDFR GPFGWEYAAV NTGELRPIDD TPEDVDMKLR QVQPLAEVRV PFTLDVTEL DSVARGATNP DLDDVARAAE RMVEAEDSAI FHGWAQAGIK GIVDSTPHEA LAVASVSDFP RAVLSAADTL R KAGVTGPY ALVLGPKAYD DLFAATQDGY PVAKQVQRLV VDGPLVRANA LAGALVMSMR GGDYELTVGQ DLSIGYAFHD RS KVELFVA ESFTFRVLEP GAAVHLRYA

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Macromolecule #2: Haliangium ochraceum Encapsulated ferritin localisation sequence

MacromoleculeName: Haliangium ochraceum Encapsulated ferritin localisation sequence
type: protein_or_peptide / ID: 2 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Haliangium ochraceum (strain DSM 14365 / JCM 11303 / SMP-2) (bacteria)
Strain: DSM 14365 / JCM 11303 / SMP-2
Molecular weightTheoretical: 14.817368 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MSSEQLHEPA ELLSEETKNM HRALVTLIEE LEAVDWYQQR ADACSEPGLH DVLIHNKNEE VEHAMMTLEW IRRRSPVFDA HMRTYLFTE RPILELEEED TGSSSSVAAS PTSAPSHGSL GIGSLRQEGK ED

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Macromolecule #3: water

MacromoleculeName: water / type: ligand / ID: 3 / Number of copies: 154 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER / Water

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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

Concentration3 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClSodium chlorideSodium Chloride
20.0 mMHEPES

Details: Solutions were prepared with MilliQ water and filtered using a 0.22 um filter.
GridModel: Quantifoil / Material: GOLD / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 281 K / Instrument: FEI VITROBOT MARK IV
Details: blot force -5, wait time 10 seconds and blot time of 3 seconds.
DetailsSample mono disperse as determined by SEC

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 11520 pixel / Digitization - Dimensions - Height: 8184 pixel / Number grids imaged: 1 / Number real images: 8109 / Average exposure time: 1.0 sec. / Average electron dose: 40.509 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 340403
CTF correctionSoftware:
Namedetails
CTFFIND (ver. 4)
RELION (ver. 3.1)CTFrefine

Details: CTF correction after motion correction followed by CFT refinement after 3D Refinement.
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1) / Software - details: Initial model / Details: Initial model produced in Relion
Final 3D classificationNumber classes: 3 / Avg.num./class: 254134 / Software - Name: RELION (ver. 3.1)
Details: Final 3D classification used to remove broken particles.
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final reconstructionNumber classes used: 1 / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 4734551
FSC plot (resolution estimation)

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Atomic model buiding 1

DetailsInitial fitting performed using chimera with real space refinement in Phenix.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Overall B value: 86 / Target criteria: CC
Output model

PDB-7oe2:
Model of closed pentamer of the Haliangium ochraceum encapsulin from symmetry expansion of icosahedral single particle reconstruction

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