+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-24002 | |||||||||
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Title | Cryo-EM map of the FLP cargo within the T. maritima encapsulin | |||||||||
Map data | Symmetry expansion and local classification of one FLP cargo protein within the T. maritima encapsulin | |||||||||
Sample |
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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 | |||||||||
Biological species | Thermotoga maritima (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.0 Å | |||||||||
Authors | LaFrance BJ / Savage DF / Nogales E | |||||||||
Funding support | United States, 2 items
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Citation | Journal: Sci Rep / Year: 2021 Title: The encapsulin from Thermotoga maritima is a flavoprotein with a symmetry matched ferritin-like cargo protein. Authors: Benjamin J LaFrance / Caleb Cassidy-Amstutz / Robert J Nichols / Luke M Oltrogge / Eva Nogales / David F Savage / Abstract: Bacterial nanocompartments, also known as encapsulins, are an emerging class of protein-based 'organelles' found in bacteria and archaea. Encapsulins are virus-like icosahedral particles comprising a ...Bacterial nanocompartments, also known as encapsulins, are an emerging class of protein-based 'organelles' found in bacteria and archaea. Encapsulins are virus-like icosahedral particles comprising a ~ 25-50 nm shell surrounding a specific cargo enzyme. Compartmentalization is thought to create a unique chemical environment to facilitate catalysis and isolate toxic intermediates. Many questions regarding nanocompartment structure-function remain unanswered, including how shell symmetry dictates cargo loading and to what extent the shell facilitates enzymatic activity. Here, we explore these questions using the model Thermotoga maritima nanocompartment known to encapsulate a redox-active ferritin-like protein. Biochemical analysis revealed the encapsulin shell to possess a flavin binding site located at the interface between capsomere subunits, suggesting the shell may play a direct and active role in the function of the encapsulated cargo. Furthermore, we used cryo-EM to show that cargo proteins use a form of symmetry-matching to facilitate encapsulation and define stoichiometry. In the case of the Thermotoga maritima encapsulin, the decameric cargo protein with fivefold symmetry preferentially binds to the pentameric-axis of the icosahedral shell. Taken together, these observations suggest the shell is not simply a passive barrier-it also plays a significant role in the structure and function of the cargo enzyme. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_24002.map.gz | 37.8 MB | EMDB map data format | |
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Header (meta data) | emd-24002-v30.xml emd-24002.xml | 16.5 KB 16.5 KB | Display Display | EMDB header |
Images | emd_24002.png | 211.2 KB | ||
Others | emd_24002_additional_1.map.gz | 49.9 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-24002 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-24002 | HTTPS FTP |
-Validation report
Summary document | emd_24002_validation.pdf.gz | 359.5 KB | Display | EMDB validaton report |
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Full document | emd_24002_full_validation.pdf.gz | 359.1 KB | Display | |
Data in XML | emd_24002_validation.xml.gz | 6.3 KB | Display | |
Data in CIF | emd_24002_validation.cif.gz | 7.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-24002 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-24002 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_24002.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Symmetry expansion and local classification of one FLP cargo protein within the T. maritima encapsulin | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.64 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Additional map: C1 reconstruction of T. maritima encapsulin showing all...
File | emd_24002_additional_1.map | ||||||||||||
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Annotation | C1 reconstruction of T. maritima encapsulin showing all FLP cargo densities present in the shell | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Cryo-EM map of FLP cargo protein within the T. maritima encapsulin
Entire | Name: Cryo-EM map of FLP cargo protein within the T. maritima encapsulin |
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Components |
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-Supramolecule #1: Cryo-EM map of FLP cargo protein within the T. maritima encapsulin
Supramolecule | Name: Cryo-EM map of FLP cargo protein within the T. maritima encapsulin type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: Thermotoga maritima (bacteria) |
Molecular weight | Theoretical: 1.83 MDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 2.0 mg/mL |
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Buffer | pH: 7.4 Details: 20 mM Tris-HCl, pH 7.4, 150 mM NaCl and 5 mM 2-mercaptoethanol |
Vitrification | Cryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: blot force. |
-Electron microscopy
Microscope | FEI TITAN |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Number grids imaged: 1 / Number real images: 1276 / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.8000000000000003 µm / Nominal defocus min: 1.5 µm |
Sample stage | Specimen holder model: GATAN 626 SINGLE TILT LIQUID NITROGEN CRYO TRANSFER HOLDER Cooling holder cryogen: NITROGEN |
+Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Chain ID: A |
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Details | The homology model for the encapsulated FLP was created using Biozentrum's SWISS-MODEL, with 5DA5 as the reference structure. This was then rigid body docked into the FLP map using ChimeraX followed by PHENIX. The final model can be found in the supplementary materials of this manuscript at 'FLP-pentamer_model.py' |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Overall B value: 110 |