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- EMDB-50586: CryoEM structure of Encapsulin::tdNfsB with an open pore state -

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Basic information

Entry
Database: EMDB / ID: EMD-50586
TitleCryoEM structure of Encapsulin::tdNfsB with an open pore state
Map data
Sample
  • Complex: Cryo-EM structure of encapsulin shell containing engineered tandem dimer NfsB
    • Protein or peptide: 29 kDa antigen Cfp29
KeywordsNanocompartment / BIOSYNTHETIC PROTEIN
Function / homologyType 1 encapsulin shell protein / Encapsulating protein for peroxidase / : / encapsulin nanocompartment / Type 1 encapsulin shell protein
Function and homology information
Biological speciesMycolicibacterium smegmatis (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.22 Å
AuthorsCapper MJ / Kohhnke J
Funding supportEuropean Union, 1 items
OrganizationGrant numberCountry
European Research Council (ERC)101002326European Union
CitationJournal: RSC Chem Biol / Year: 2024
Title: A nanoengineered tandem nitroreductase: designing a robust prodrug-activating nanoreactor.
Authors: Mariia Zmyslia / Michael J Capper / Michael Grimmeisen / Kerstin Sartory / Benedikt Deuringer / Mohamed Abdelsalam / Kaiwei Shen / Manfred Jung / Wolfgang Sippl / Hans-Georg Koch / Laurine ...Authors: Mariia Zmyslia / Michael J Capper / Michael Grimmeisen / Kerstin Sartory / Benedikt Deuringer / Mohamed Abdelsalam / Kaiwei Shen / Manfred Jung / Wolfgang Sippl / Hans-Georg Koch / Laurine Kaul / Regine Süss / Jesko Köhnke / Claudia Jessen-Trefzer /
Abstract: Nitroreductases are important enzymes for a variety of applications, including cancer therapy and bioremediation. They often require encapsulation to improve stability and activity. We focus on ...Nitroreductases are important enzymes for a variety of applications, including cancer therapy and bioremediation. They often require encapsulation to improve stability and activity. We focus on genetically encoded encapsulation of nitroreductases within protein capsids, like encapsulins. Our study showcases the encapsulation of nitroreductase NfsB as functional dimers within encapsulins, which enhances protein activity and stability in diverse conditions. Mutations within the pore region are beneficial for activity of the encapsulated enzyme, potentially by increasing diffusion rates. Cryogenic electron microscopy reveals the overall architecture of the encapsulated dimeric NfsB within the nanoreactor environment and identifies multiple pore states in the shell. These findings highlight the potential of encapsulins as versatile tools for enhancing enzyme performance across various fields.
History
DepositionJun 9, 2024-
Header (metadata) releaseMar 19, 2025-
Map releaseMar 19, 2025-
UpdateMar 19, 2025-
Current statusMar 19, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_50586.png

Downloads & links

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Map

FileDownload / File: emd_50586.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

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AxesZ (Sec.)Y (Row.)X (Col.)
0.78 Å/pix.
x 512 pix.
= 400.998 Å		0.78 Å/pix.
x 512 pix.
= 400.998 Å		0.78 Å/pix.
x 512 pix.
= 400.998 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.7832 Å
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Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.037
Minimum - Maximum-0.03807873 - 0.17778113
Average (Standard dev.)0.00046643714 (±0.009696989)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 400.9984 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_50586_msk_1.map
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Half map: #2

Fileemd_50586_half_map_1.map
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Half map: #1

Fileemd_50586_half_map_2.map
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Sample components

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Entire : Cryo-EM structure of encapsulin shell containing engineered tande...

EntireName: Cryo-EM structure of encapsulin shell containing engineered tandem dimer NfsB
Components
  • Complex: Cryo-EM structure of encapsulin shell containing engineered tandem dimer NfsB
    • Protein or peptide: 29 kDa antigen Cfp29

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Supramolecule #1: Cryo-EM structure of encapsulin shell containing engineered tande...

SupramoleculeName: Cryo-EM structure of encapsulin shell containing engineered tandem dimer NfsB
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Mycolicibacterium smegmatis (bacteria)

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Macromolecule #1: 29 kDa antigen Cfp29

MacromoleculeName: 29 kDa antigen Cfp29 / type: protein_or_peptide / ID: 1 / Number of copies: 60 / Enantiomer: LEVO
Source (natural)Organism: Mycolicibacterium smegmatis (bacteria)
Molecular weightTheoretical: 30.045578 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MNNLYRDLAP ITESAWAEIE LEATRTFKRH IAGRRVVDVS GPNGPTTASV STGHLLDVSP PGDGVIAHLR DAKPLVRLRV PFTVARRDI DDVERGSQDS DWDPVKDAAK KLAFVEDRAI FEGYAAASIE GIRSSSSNPA LALPDDAREI PDVIAQALSE L RLAGVDGP ...String:
MNNLYRDLAP ITESAWAEIE LEATRTFKRH IAGRRVVDVS GPNGPTTASV STGHLLDVSP PGDGVIAHLR DAKPLVRLRV PFTVARRDI DDVERGSQDS DWDPVKDAAK KLAFVEDRAI FEGYAAASIE GIRSSSSNPA LALPDDAREI PDVIAQALSE L RLAGVDGP YSVLLSAETY TKVSETTAHG YPIREHINRL VDGEIIWAPA IDGAFVLSTR GGDFDLQLGT DVSIGYLSHD AE VVHLYME ETMTFLCYTA EASVALTPEL WSHPQFEK

UniProtKB: Type 1 encapsulin shell 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

Concentration20 mg/mL
BufferpH: 7.5 / Component:
ConcentrationName
20.0 mMHEPES
100.0 mMNaCl
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 281 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Specialist opticsEnergy filter - Name: In-column Omega Filter
Image recordingFilm or detector model: DIRECT ELECTRON APOLLO (4k x 4k) / Number grids imaged: 1 / Average exposure time: 1.5 sec. / Average electron dose: 60.2 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 80000
Sample stageSpecimen holder model: JEOL / Cooling holder cryogen: NITROGEN

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

7boj

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: I (icosahedral) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.22 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 546841
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC
Final angle assignmentType: ANGULAR RECONSTITUTION
Final 3D classificationNumber classes: 3
FSC plot (resolution estimation)

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