+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-13670 | ||||||||||||||||||
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Title | isolated S-layer of Ca.M.Lanthanididphila | ||||||||||||||||||
Map data | Subtomogram average of the Ca.M.lanthanidiphila S-layer obtained from cryo-tomograms of isolated S-layer patches. 6-fold rotational symmetry has been applied. | ||||||||||||||||||
Sample |
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Keywords | S-layer / STRUCTURAL PROTEIN | ||||||||||||||||||
Biological species | Candidatus Methylomirabilis lanthanidiphila (bacteria) | ||||||||||||||||||
Method | subtomogram averaging / cryo EM / Resolution: 21.0 Å | ||||||||||||||||||
Authors | Gambelli L / Mesman R | ||||||||||||||||||
Funding support | 5 items
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Citation | Journal: Front Microbiol / Year: 2021 Title: The Polygonal Cell Shape and Surface Protein Layer of Anaerobic Methane-Oxidizing Bacteria. Authors: Lavinia Gambelli / Rob Mesman / Wouter Versantvoort / Christoph A Diebolder / Andreas Engel / Wiel Evers / Mike S M Jetten / Martin Pabst / Bertram Daum / Laura van Niftrik / Abstract: bacteria perform anaerobic methane oxidation coupled to nitrite reduction via an intra-aerobic pathway, producing carbon dioxide and dinitrogen gas. These diderm bacteria possess an unusual ... bacteria perform anaerobic methane oxidation coupled to nitrite reduction via an intra-aerobic pathway, producing carbon dioxide and dinitrogen gas. These diderm bacteria possess an unusual polygonal cell shape with sharp ridges that run along the cell body. Previously, a putative surface protein layer (S-layer) was observed as the outermost cell layer of these bacteria. We hypothesized that this S-layer is the determining factor for their polygonal cell shape. Therefore, we enriched the S-layer from cells and through LC-MS/MS identified a 31 kDa candidate S-layer protein, mela_00855, which had no homology to any other known protein. Antibodies were generated against a synthesized peptide derived from the mela_00855 protein sequence and used in immunogold localization to verify its identity and location. Both on thin sections of cells and in negative-stained enriched S-layer patches, the immunogold localization identified mela_00855 as the S-layer protein. Using electron cryo-tomography and sub-tomogram averaging of S-layer patches, we observed that the S-layer has a hexagonal symmetry. Cryo-tomography of whole cells showed that the S-layer and the outer membrane, but not the peptidoglycan layer and the cytoplasmic membrane, exhibited the polygonal shape. Moreover, the S-layer consisted of multiple rigid sheets that partially overlapped, most likely giving rise to the unique polygonal cell shape. These characteristics make the S-layer of a distinctive and intriguing case to study. | ||||||||||||||||||
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_13670.map.gz | 46.8 MB | EMDB map data format | |
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Header (meta data) | emd-13670-v30.xml emd-13670.xml | 11.9 KB 11.9 KB | Display Display | EMDB header |
Images | emd_13670.png | 188.6 KB | ||
Filedesc metadata | emd-13670.cif.gz | 4.4 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-13670 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13670 | HTTPS FTP |
-Validation report
Summary document | emd_13670_validation.pdf.gz | 666.7 KB | Display | EMDB validaton report |
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Full document | emd_13670_full_validation.pdf.gz | 666.2 KB | Display | |
Data in XML | emd_13670_validation.xml.gz | 6.6 KB | Display | |
Data in CIF | emd_13670_validation.cif.gz | 7.4 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13670 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13670 | HTTPS FTP |
-Related structure data
Related structure data | C: citing same article (ref.) |
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Similar structure data | |
EM raw data | EMPIAR-10822 (Title: Cryo Electron Tomography of isolated S-layer patches from Ca.M.lanthanidiphila Data size: 46.3 Data #1: Reconstructed tomogram of isolated Ca. M.lanthanidiphila S-layer patches [reconstructed volumes] Data #2: Reconstructed tomogram of isolated Ca. M.lanthanidiphila S-layer patches [reconstructed volumes]) |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_13670.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Subtomogram average of the Ca.M.lanthanidiphila S-layer obtained from cryo-tomograms of isolated S-layer patches. 6-fold rotational symmetry has been applied. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.62631 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : isolated S-layer of Ca.M.lanthanidiphila
Entire | Name: isolated S-layer of Ca.M.lanthanidiphila |
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Components |
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-Supramolecule #1: isolated S-layer of Ca.M.lanthanidiphila
Supramolecule | Name: isolated S-layer of Ca.M.lanthanidiphila / type: organelle_or_cellular_component / ID: 1 / Parent: 0 Details: S-layer was isolated by boiling disrupted flocks of Ca.M.Lanthanidiphila enrichment culture in 4% SDS. |
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Source (natural) | Organism: Candidatus Methylomirabilis lanthanidiphila (bacteria) Location in cell: on top of outer membrane |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | subtomogram averaging |
Aggregation state | 2D array |
-Sample preparation
Buffer | pH: 7 |
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Grid | Model: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 40 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 20.0 kPa / Details: glow-discharged at 15 mA current |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV Details: 2 microlitre sample was mixed with 0.5 microlitre 10 nm ProteinA gold solution Grids were plunge frozen in liquid ethane, blot force 2, blot time 2.5-3 sec.. |
Details | concentrated isolated S-layer in ultrapure water |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Temperature | Max: 93.0 K |
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Average exposure time: 0.3 sec. / Average electron dose: 1.71538 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Calibrated defocus max: 4.7 µm / Calibrated defocus min: 4.43 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 4.0 µm / Nominal magnification: 33000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Number classes used: 1 / Applied symmetry - Point group: C6 (6 fold cyclic) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 21.0 Å / Resolution method: OTHER / Software - Name: PEET (ver. 1.14.0) Details: Resolution of the Ca. M. lanthanidiphila S-layer sub-tomogram average was determined by FFT Number subtomograms used: 8938 |
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Extraction | Number tomograms: 2 / Number images used: 11274 / Method: random grid / Software - Name: PEET (ver. 1.14.0) |
Final angle assignment | Type: NOT APPLICABLE |