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- EMDB-23384: Thermotoga maritima Encapsulin Nanocompartment Pore Mutant S5D -

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Basic information

Entry
Database: EMDB / ID: EMD-23384
TitleThermotoga maritima Encapsulin Nanocompartment Pore Mutant S5D
Map data
Sample
  • Complex: Thermotoga maritima Encapsulin Nanocompartment Pore Mutant S5D
    • Protein or peptide: Maritimacin
  • Ligand: RIBOFLAVIN
KeywordsEncapsulin / Nanocompartment / VIRUS LIKE PARTICLE
Function / homology
Function and homology information


encapsulin nanocompartment / Hydrolases; Acting on peptide bonds (peptidases) / peptidase activity / iron ion transport / intracellular iron ion homeostasis / proteolysis
Similarity search - Function
Type 1 encapsulin shell protein / Encapsulating protein for peroxidase / :
Similarity search - Domain/homology
Type 1 encapsulin shell protein
Similarity search - Component
Biological speciesThermotoga maritima MSB8 (bacteria) / Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.96 Å
AuthorsAndreas MP / Adamson L
Funding support Australia, United States, 3 items
OrganizationGrant numberCountry
Australian Research Council (ARC)DE190100624 Australia
Other privateWRF2020 Australia
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM133325 United States
CitationJournal: Sci Adv / Year: 2022
Title: Pore structure controls stability and molecular flux in engineered protein cages.
Authors: Lachlan S R Adamson / Nuren Tasneem / Michael P Andreas / William Close / Eric N Jenner / Taylor N Szyszka / Reginald Young / Li Chen Cheah / Alexander Norman / Hugo I MacDermott-Opeskin / ...Authors: Lachlan S R Adamson / Nuren Tasneem / Michael P Andreas / William Close / Eric N Jenner / Taylor N Szyszka / Reginald Young / Li Chen Cheah / Alexander Norman / Hugo I MacDermott-Opeskin / Megan L O'Mara / Frank Sainsbury / Tobias W Giessen / Yu Heng Lau /
Abstract: Protein cages are a common architectural motif used by living organisms to compartmentalize and control biochemical reactions. While engineered protein cages have featured in the construction of ...Protein cages are a common architectural motif used by living organisms to compartmentalize and control biochemical reactions. While engineered protein cages have featured in the construction of nanoreactors and synthetic organelles, relatively little is known about the underlying molecular parameters that govern stability and flux through their pores. In this work, we systematically designed 24 variants of the encapsulin cage, featuring pores of different sizes and charges. Twelve pore variants were successfully assembled and purified, including eight designs with exceptional thermal stability. While negatively charged mutations were better tolerated, we were able to form stable assemblies covering a full range of pore sizes and charges, as observed in seven new cryo-EM structures at 2.5- to 3.6-Å resolution. Molecular dynamics simulations and stopped-flow experiments revealed the importance of considering both pore size and charge, together with flexibility and rate-determining steps, when designing protein cages for controlling molecular flux.
History
DepositionJan 27, 2021-
Header (metadata) releaseFeb 9, 2022-
Map releaseFeb 9, 2022-
UpdateMay 29, 2024-
Current statusMay 29, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.6
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.6
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7lis
  • Surface level: 0.6
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7lis
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23384.png

Downloads & links

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Map

FileDownload / File: emd_23384.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.99 Å/pix.
x 384 pix.
= 378.624 Å		0.99 Å/pix.
x 384 pix.
= 378.624 Å		0.99 Å/pix.
x 384 pix.
= 378.624 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.986 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.6 / Movie #1: 0.6
Minimum - Maximum-2.0525596 - 3.4106612
Average (Standard dev.)0.003352073 (±0.19071737)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 378.624 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.9860.9860.986
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z378.624378.624378.624
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS384384384
D min/max/mean-2.0533.4110.003

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Supplemental data

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Sample components

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Entire : Thermotoga maritima Encapsulin Nanocompartment Pore Mutant S5D

EntireName: Thermotoga maritima Encapsulin Nanocompartment Pore Mutant S5D
Components
  • Complex: Thermotoga maritima Encapsulin Nanocompartment Pore Mutant S5D
    • Protein or peptide: Maritimacin
  • Ligand: RIBOFLAVIN

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Supramolecule #1: Thermotoga maritima Encapsulin Nanocompartment Pore Mutant S5D

SupramoleculeName: Thermotoga maritima Encapsulin Nanocompartment Pore Mutant S5D
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Thermotoga maratima encapsulin pore mutant with E184D, P189D, and deletion of amino acids A185-Y188
Source (natural)Organism: Thermotoga maritima MSB8 (bacteria)
Molecular weightTheoretical: 1.825782 MDa

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Macromolecule #1: Maritimacin

MacromoleculeName: Maritimacin / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: Hydrolases; Acting on peptide bonds (peptidases)
Source (natural)Organism: Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099) (bacteria)
Strain: ATCC 43589 / MSB8 / DSM 3109 / JCM 10099
Molecular weightTheoretical: 30.091287 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MEFLKRSFAP LTEKQWQEID NRAREIFKTQ LYGRKFVDVE GPYGWEYAAH PLGEVEVLSD ENEVVKWGLR KSLPLIELRA TFTLDLWEL DNLERGKPNV DLSSLEETVR KVAEFEDEVI FRGCEKSGVK GLLSFEERKI ECGSTPKDLL EAIVRALSIF S KDGIEGPY ...String:
MEFLKRSFAP LTEKQWQEID NRAREIFKTQ LYGRKFVDVE GPYGWEYAAH PLGEVEVLSD ENEVVKWGLR KSLPLIELRA TFTLDLWEL DNLERGKPNV DLSSLEETVR KVAEFEDEVI FRGCEKSGVK GLLSFEERKI ECGSTPKDLL EAIVRALSIF S KDGIEGPY TLVINTDRWI NFLKEDDLEK RVEECLRGGK IITTPRIEDA LVVSERGGDF KLILGQDLSI GYEDREKDAV RL FITETFT FQVVNPEALI LLKF

UniProtKB: Type 1 encapsulin shell protein

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Macromolecule #2: RIBOFLAVIN

MacromoleculeName: RIBOFLAVIN / type: ligand / ID: 2 / Number of copies: 1 / Formula: RBF
Molecular weightTheoretical: 376.364 Da
Chemical component information

ChemComp-RBF:
RIBOFLAVIN

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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

Concentration2.0 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
100.0 mMNaClSodium Chloride
20.0 mMC4H11NO3Tris
GridModel: C-flat-2/2 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: blot force 0, wait time 30 seconds.

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Electron microscopy

MicroscopeTFS TALOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Average exposure time: 47.84 sec. / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: -2.5 µm / Nominal defocus min: -1.0 µm
Sample stageCooling holder cryogen: NITROGEN

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Image processing

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

3dkt


Details: Initial map for refinements made from PDB entry 3DKT
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.96 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.15.0) / Number images used: 33416
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 2.15.0)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 2.15.0)

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Atomic model buiding 1

Initial modelPDB ID:

3dkt


Chain - Chain ID: A / Chain - Residue range: 1-265 / Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 51.86
Output model

PDB-7lis:
Thermotoga maritima Encapsulin Nanocompartment Pore Mutant S5D

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