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- EMDB-27654: A subtomogram average of H. neapolitanus Rubisco within alpha-car... -

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Basic information

Entry
Database: EMDB / ID: EMD-27654
TitleA subtomogram average of H. neapolitanus Rubisco within alpha-carboxysomes
Map dataSubtomogram average of Rubisco within H. neapolitanus alpha-carboxysome. Contour level for best display of interior.
Sample
  • Organelle or cellular component: Rubisco within the alpha-carboxysome
KeywordsCarbon fixation / alpha-carboxysome / enzyme / LYASE
Biological speciesHalothiobacillus neapolitanus (bacteria)
Methodsubtomogram averaging / cryo EM / Resolution: 4.5 Å
AuthorsMetskas LA / Blikstad C / Laughlin T / Savage DF / Jensen GJ
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1F32GM135994-01 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01AI127401 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM129241 United States
CitationJournal: Nat Commun / Year: 2022
Title: Rubisco forms a lattice inside alpha-carboxysomes.
Authors: Lauren Ann Metskas / Davi Ortega / Luke M Oltrogge / Cecilia Blikstad / Derik R Lovejoy / Thomas G Laughlin / David F Savage / Grant J Jensen /
Abstract: Despite the importance of microcompartments in prokaryotic biology and bioengineering, structural heterogeneity has prevented a complete understanding of their architecture, ultrastructure, and ...Despite the importance of microcompartments in prokaryotic biology and bioengineering, structural heterogeneity has prevented a complete understanding of their architecture, ultrastructure, and spatial organization. Here, we employ cryo-electron tomography to image α-carboxysomes, a pseudo-icosahedral microcompartment responsible for carbon fixation. We have solved a high-resolution subtomogram average of the Rubisco cargo inside the carboxysome, and determined the arrangement of the enzyme. We find that the H. neapolitanus Rubisco polymerizes in vivo, mediated by the small Rubisco subunit. These fibrils can further pack to form a lattice with six-fold pseudo-symmetry. This arrangement preserves freedom of motion and accessibility around the Rubisco active site and the binding sites for two other carboxysome proteins, CsoSCA (a carbonic anhydrase) and the disordered CsoS2, even at Rubisco concentrations exceeding 800 μM. This characterization of Rubisco cargo inside the α-carboxysome provides insight into the balance between order and disorder in microcompartment organization.
History
DepositionJul 19, 2022-
Header (metadata) releaseAug 3, 2022-
Map releaseAug 3, 2022-
UpdateJan 17, 2024-
Current statusJan 17, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27654.png

Downloads & links

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Map

FileDownload / File: emd_27654.map.gz / Format: CCP4 / Size: 15.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSubtomogram average of Rubisco within H. neapolitanus alpha-carboxysome. Contour level for best display of interior.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 160 pix.
= 176.64 Å		1.1 Å/pix.
x 160 pix.
= 176.64 Å		1.1 Å/pix.
x 160 pix.
= 176.64 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.104 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.588
Minimum - Maximum-6.1842604 - 5.3301263
Average (Standard dev.)0.048541438 (±0.7444289)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions160160160
Spacing160160160
CellA=B=C: 176.64 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_27654_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half-map 1, unfiltered

Fileemd_27654_half_map_1.map
AnnotationHalf-map 1, unfiltered
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half-map 2, unfiltered

Fileemd_27654_half_map_2.map
AnnotationHalf-map 2, unfiltered
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Rubisco within the alpha-carboxysome

EntireName: Rubisco within the alpha-carboxysome
Components
  • Organelle or cellular component: Rubisco within the alpha-carboxysome

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Supramolecule #1: Rubisco within the alpha-carboxysome

SupramoleculeName: Rubisco within the alpha-carboxysome / type: organelle_or_cellular_component / ID: 1 / Parent: 0
Details: Subtomogram average of Rubisco within carboxysomes purified from H. neapolitanus
Source (natural)Organism: Halothiobacillus neapolitanus (bacteria) / Location in cell: Alpha-carboxysome

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferpH: 8
VitrificationCryogen name: ETHANE
DetailsPurified alpha-carboxysome

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Average electron dose: 2.9 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 81000
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.5 Å / Resolution method: FSC 0.143 CUT-OFF
Software:
Namedetails
Dynamo (ver. 1.1.333)Final subtomogram average
RELION (ver. 3.1)Final post-processed map

Number subtomograms used: 24698
ExtractionNumber tomograms: 62 / Number images used: 32930
Final angle assignmentType: ANGULAR RECONSTITUTION / Details: weighted back projection
FSC plot (resolution estimation)

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