+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-10388 | |||||||||
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Title | In situ structure of the Caulobacter crescentus S-layer | |||||||||
Map data | In situ structure of the Caulobacter crescentus S-layer | |||||||||
Sample |
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Function / homology | RsaA N-terminal domain / S-layer / RTX calcium-binding nonapeptide repeat / RTX calcium-binding nonapeptide repeat (4 copies) / Serralysin-like metalloprotease, C-terminal / calcium ion binding / extracellular region / S-layer protein Function and homology information | |||||||||
Biological species | Caulobacter crescentus NA1000 (bacteria) | |||||||||
Method | subtomogram averaging / cryo EM / Resolution: 4.8 Å | |||||||||
Authors | Bharat T / von Kuegelgen A | |||||||||
Funding support | United Kingdom, 1 items
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Citation | Journal: Cell / Year: 2020 Title: In Situ Structure of an Intact Lipopolysaccharide-Bound Bacterial Surface Layer. Authors: Andriko von Kügelgen / Haiping Tang / Gail G Hardy / Danguole Kureisaite-Ciziene / Yves V Brun / Phillip J Stansfeld / Carol V Robinson / Tanmay A M Bharat / Abstract: Most bacterial and all archaeal cells are encapsulated by a paracrystalline, protective, and cell-shape-determining proteinaceous surface layer (S-layer). On Gram-negative bacteria, S-layers are ...Most bacterial and all archaeal cells are encapsulated by a paracrystalline, protective, and cell-shape-determining proteinaceous surface layer (S-layer). On Gram-negative bacteria, S-layers are anchored to cells via lipopolysaccharide. Here, we report an electron cryomicroscopy structure of the Caulobacter crescentus S-layer bound to the O-antigen of lipopolysaccharide. Using native mass spectrometry and molecular dynamics simulations, we deduce the length of the O-antigen on cells and show how lipopolysaccharide binding and S-layer assembly is regulated by calcium. Finally, we present a near-atomic resolution in situ structure of the complete S-layer using cellular electron cryotomography, showing S-layer arrangement at the tip of the O-antigen. A complete atomic structure of the S-layer shows the power of cellular tomography for in situ structural biology and sheds light on a very abundant class of self-assembling molecules with important roles in prokaryotic physiology with marked potential for synthetic biology and surface-display applications. | |||||||||
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_10388.map.gz | 28.2 MB | EMDB map data format | |
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Header (meta data) | emd-10388-v30.xml emd-10388.xml | 12.7 KB 12.7 KB | Display Display | EMDB header |
Images | emd_10388.png | 121.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-10388 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-10388 | HTTPS FTP |
-Related structure data
Related structure data | 6z7pMC 6t72C M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_10388.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | In situ structure of the Caulobacter crescentus S-layer | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.35 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 : Caulobacter crescentus S-layer
Entire | Name: Caulobacter crescentus S-layer |
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Components |
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-Supramolecule #1: Caulobacter crescentus S-layer
Supramolecule | Name: Caulobacter crescentus S-layer / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: Caulobacter crescentus S-layer |
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Source (natural) | Organism: Caulobacter crescentus NA1000 (bacteria) / Strain: YB2811 / Location in cell: extra-cellular |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | subtomogram averaging |
Aggregation state | cell |
-Sample preparation
Buffer | pH: 7 / Details: PYE medium |
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Grid | Model: Quantifoil R2/2 / Material: COPPER/RHODIUM / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Details: 15 mA |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283.15 K / Instrument: FEI VITROBOT MARK IV / Details: 1.5 s blot. |
Details | Caulobacter crescentus stalk |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Calibrated defocus max: -5.0 µm / Calibrated defocus min: -2.0 µm / Calibrated magnification: 105000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: -5.0 µm / Nominal defocus min: -2.0 µm / Nominal magnification: 105000 |
Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Temperature | Min: 70.0 K / Max: 70.0 K |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Average exposure time: 1.0 sec. / Average electron dose: 3.4 e/Å2 / Details: Dose symmetric tilt scheme (Hagen et al, JSB) |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Extraction | Number tomograms: 110 / Number images used: 51866 / Reference model: Ab initio / Method: RELION / Software - Name: RELION (ver. 1.4) / Details: RELION subtomogram averaging |
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CTF correction | Details: Following Turonova and Briggs NovaCTF |
Final 3D classification | Number classes: 1 / Avg.num./class: 1 / Details: No classification done |
Final angle assignment | Type: OTHER / Details: Using the AV3 package (Foerster et al, PNAS 2006) |
Final reconstruction | Number classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 4.8 Å / Resolution method: FSC 0.143 CUT-OFF Details: Tomogram generation in IMOD using the NovaCTF implementation of Turonova and Briggs. Sub-tomogram averaging performed using the AV3 package (Friedrich Foerster) applied to tubular specimens ...Details: Tomogram generation in IMOD using the NovaCTF implementation of Turonova and Briggs. Sub-tomogram averaging performed using the AV3 package (Friedrich Foerster) applied to tubular specimens (Bharat et al, PLoS Biology, 2011) Number subtomograms used: 51866 |
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: Cross-correlation coefficient |
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Output model | PDB-6z7p: |