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- EMDB-22094: CryoEM structure of the holo-SrpI encapsulin complex from Synecho... -

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Basic information

Entry
Database: EMDB / ID: EMD-22094
TitleCryoEM structure of the holo-SrpI encapsulin complex from Synechococcus elongatus PCC 7942
Map dataCryo-EM Structure of the holo-Srpl encapsulin complex from Synechococcus elongatus PCC 7942
Sample
  • Complex: SrpI encapsulin holo complex
    • Complex: SrpI encapsulin
      • Protein or peptide: Protein SrpI
    • Complex: CyD cargo protein
Keywordsencapsulin / nanocompartment / cysteine desulfurase / HK97-fold / VIRUS LIKE PARTICLE
Function / homologyType 2A encapsulin shell protein SrpI-like / : / Type 2A encapsulin shell protein SrpI-like / encapsulin nanocompartment / Type 2A encapsulin shell protein SrpI
Function and homology information
Biological speciesSynechococcus elongatus (strain PCC 7942 / FACHB-805) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.2 Å
AuthorsLaFrance BJ / Nichols RJ / Phillips NR / Oltrogge LM / Valentin-Alvarado LE / Bischoff AJ / Savage DF / Nogales E
Funding support United States, 2 items
OrganizationGrant numberCountry
Department of Energy (DOE, United States)DE- 638 SC00016240 United States
National Science Foundation (NSF, United States)1106400 United States
Citation
Journal: Elife / Year: 2021
Title: Discovery and characterization of a novel family of prokaryotic nanocompartments involved in sulfur metabolism.
Authors: Robert J Nichols / Benjamin LaFrance / Naiya R Phillips / Devon R Radford / Luke M Oltrogge / Luis E Valentin-Alvarado / Amanda J Bischoff / Eva Nogales / David F Savage /
Abstract: Prokaryotic nanocompartments, also known as encapsulins, are a recently discovered proteinaceous organelle-like compartment in prokaryotes that compartmentalize cargo enzymes. While initial studies ...Prokaryotic nanocompartments, also known as encapsulins, are a recently discovered proteinaceous organelle-like compartment in prokaryotes that compartmentalize cargo enzymes. While initial studies have begun to elucidate the structure and physiological roles of encapsulins, bioinformatic evidence suggests that a great diversity of encapsulin nanocompartments remains unexplored. Here, we describe a novel encapsulin in the freshwater cyanobacterium PCC 7942. This nanocompartment is upregulated upon sulfate starvation and encapsulates a cysteine desulfurase enzyme via an N-terminal targeting sequence. Using cryo-electron microscopy, we have determined the structure of the nanocompartment complex to 2.2 Å resolution. Lastly, biochemical characterization of the complex demonstrated that the activity of the cysteine desulfurase is enhanced upon encapsulation. Taken together, our discovery, structural analysis, and enzymatic characterization of this prokaryotic nanocompartment provide a foundation for future studies seeking to understand the physiological role of this encapsulin in various bacteria.
#1: Journal: Biorxiv / Year: 2020
Title: Discovery and characterization of a novel family of prokaryotic nanocompartments involved in sulfur metabolism
Authors: Nichols RJ / LaFrance BJ / Phillips NR / Oltrogge LM / Valentin-Alvarado LE / Bischoff AJ / Nogales E / Savage DF
History
DepositionJun 1, 2020-
Header (metadata) releaseJun 10, 2020-
Map releaseJun 10, 2020-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6x8m
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6x8m
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22094.png

Downloads & links

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Map

FileDownload / File: emd_22094.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM Structure of the holo-Srpl encapsulin complex from Synechococcus elongatus PCC 7942
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.86 Å/pix.
x 512 pix.
= 441.6 Å		0.86 Å/pix.
x 512 pix.
= 441.6 Å		0.86 Å/pix.
x 512 pix.
= 441.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.8625 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.02 / Movie #1: 0.02
Minimum - Maximum-0.05023677 - 0.10099724
Average (Standard dev.)0.00013741106 (±0.0034197264)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 441.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.86250.86250.8625
M x/y/z512512512
origin x/y/z0.0000.0000.000
length x/y/z441.600441.600441.600
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS512512512
D min/max/mean-0.0500.1010.000

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Supplemental data

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Mask #1

Fileemd_22094_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Even half-map for holo-Srpl encapsulin complex

Fileemd_22094_half_map_1.map
AnnotationEven half-map for holo-Srpl encapsulin complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Odd half-map for holo-Srpl encapsulin complex

Fileemd_22094_half_map_2.map
AnnotationOdd half-map for holo-Srpl encapsulin complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : SrpI encapsulin holo complex

EntireName: SrpI encapsulin holo complex
Components
  • Complex: SrpI encapsulin holo complex
    • Complex: SrpI encapsulin
      • Protein or peptide: Protein SrpI
    • Complex: CyD cargo protein

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Supramolecule #1: SrpI encapsulin holo complex

SupramoleculeName: SrpI encapsulin holo complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Molecular weightTheoretical: 2.1 MDa

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Supramolecule #2: SrpI encapsulin

SupramoleculeName: SrpI encapsulin / type: complex / ID: 2 / Parent: 1 / Macromolecule list: all
Source (natural)Organism: Synechococcus elongatus (strain PCC 7942 / FACHB-805) (bacteria)

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Supramolecule #3: CyD cargo protein

SupramoleculeName: CyD cargo protein / type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Synechococcus elongatus (strain PCC 7942 / FACHB-805) (bacteria)

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Macromolecule #1: Protein SrpI

MacromoleculeName: Protein SrpI / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Synechococcus elongatus (strain PCC 7942 / FACHB-805) (bacteria)
Strain: PCC 7942 / FACHB-805
Molecular weightTheoretical: 35.371121 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MTDNAPQLAL RDVAARQLAN ATKTVPQLRT ITPRWLVRLL HWTPVEAGIY RVNQVKDASQ ITVACSERDE SELPETFVDY IDNPREYLL SAVNTVVDVH TRISDLYSNP HDQIREQLRL TIEIMKERQE SELINSREYG LLNNVAPGQL VHTRNGAPTP D DLDELLIR ...String:
MTDNAPQLAL RDVAARQLAN ATKTVPQLRT ITPRWLVRLL HWTPVEAGIY RVNQVKDASQ ITVACSERDE SELPETFVDY IDNPREYLL SAVNTVVDVH TRISDLYSNP HDQIREQLRL TIEIMKERQE SELINSREYG LLNNVAPGQL VHTRNGAPTP D DLDELLIR VWKEPAFFLA HPQAIAAFGR ECTRRGVPPA TVSLFGSSFI TWRGVPLIPS DKVPLENGKT KILLLRVGES RQ GVVGLYQ PNLPGEQGMG LSVRFMGINR KALASYLVSL YCSLAVLTDD ALAVLDNVDV TQYHTYRYNS GHHHHHH

UniProtKB: Type 2A encapsulin shell protein SrpI

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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

Concentration2.6 mg/mL
BufferpH: 10
GridModel: Quantifoil, UltrAuFoil, R1.2/1.3 / Material: GOLD / Mesh: 300
Details: Open hole grids were treated with 2 drops of chloroform, and were not glow discharged before applying sample.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 278 K / Instrument: FEI VITROBOT MARK IV
Details: blot force 5, 3 sec blot, 100 humidity, 4C, 1 sec drain time.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 25 eV
SoftwareName: SerialEM
DetailsResidual beam tilt corrected in RELION/3.1. CTF refinement performed in RELION/3.1.
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Number real images: 1027 / Average electron dose: 35.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.6 mm / Nominal defocus max: 2.8000000000000003 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 64000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsImages collected in super-resolution mode
Particle selectionNumber selected: 15686 / Details: Relion LoG autopicker to select particles
Startup modelType of model: OTHER
Details: Initial reference model was the negative stain structure low pass filtered to 30A. Once the refinement was better than 3.5A, then a 5A map was used as the reference model to enforce proper handedness.
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 2.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 11980
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

SoftwareName: PHENIX (ver. dev-2883)
DetailsOne subunit was built completely de novo based on sequence/density, and iteratively real space refined with neighboring subunits using PHENIX.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Overall B value: 20 / Target criteria: Correlation coefficient
Output model

PDB-6x8m:
CryoEM structure of the holo-SrpI encapsulin complex from Synechococcus elongatus PCC 7942

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