+Open data
-Basic information
Entry | Database: PDB / ID: 5n97 | ||||||||||||
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Title | Structure of the C. crescentus S-layer | ||||||||||||
Components | S-layer protein rsaA | ||||||||||||
Keywords | STRUCTURAL PROTEIN / S-layer / sub-tomogram averaging / bacteria / cell surface / caulobacter | ||||||||||||
Function / homology | RsaA N-terminal domain / RTX calcium-binding nonapeptide repeat / RTX calcium-binding nonapeptide repeat (4 copies) / Serralysin-like metalloprotease, C-terminal / calcium ion binding / S-layer protein rsaA Function and homology information | ||||||||||||
Biological species | Caulobacter crescentus NA1000 (bacteria) | ||||||||||||
Method | ELECTRON MICROSCOPY / subtomogram averaging / cryo EM / Resolution: 7.4 Å | ||||||||||||
Authors | Bharat, T.A. / Hagen, W.J. / Briggs, J.A. / Lowe, J. | ||||||||||||
Funding support | United Kingdom, Germany, 3items
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Citation | Journal: Nat Microbiol / Year: 2017 Title: Structure of the hexagonal surface layer on Caulobacter crescentus cells. Authors: Tanmay A M Bharat / Danguole Kureisaite-Ciziene / Gail G Hardy / Ellen W Yu / Jessica M Devant / Wim J H Hagen / Yves V Brun / John A G Briggs / Jan Löwe / Abstract: Many prokaryotic cells are encapsulated by a surface layer (S-layer) consisting of repeating units of S-layer proteins. S-layer proteins are a diverse class of molecules found in Gram-positive and ...Many prokaryotic cells are encapsulated by a surface layer (S-layer) consisting of repeating units of S-layer proteins. S-layer proteins are a diverse class of molecules found in Gram-positive and Gram-negative bacteria and most archaea. S-layers protect cells from the outside, provide mechanical stability and also play roles in pathogenicity. In situ structural information about this highly abundant class of proteins is scarce, so atomic details of how S-layers are arranged on the surface of cells have remained elusive. Here, using purified Caulobacter crescentus' sole S-layer protein RsaA, we obtained a 2.7 Å X-ray structure that shows the hexameric S-layer lattice. We also solved a 7.4 Å structure of the S-layer through electron cryotomography and sub-tomogram averaging of cell stalks. The X-ray structure was docked unambiguously into the electron cryotomography map, resulting in a pseudo-atomic-level description of the in vivo S-layer, which agrees completely with the atomic X-ray lattice model. The cellular S-layer atomic structure shows that the S-layer is porous, with a largest gap dimension of 27 Å, and is stabilized by multiple Ca ions bound near the interfaces. This study spans different spatial scales from atoms to cells by combining X-ray crystallography with electron cryotomography and sub-nanometre-resolution sub-tomogram averaging. | ||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 5n97.cif.gz | 762.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb5n97.ent.gz | 637.5 KB | Display | PDB format |
PDBx/mmJSON format | 5n97.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/n9/5n97 ftp://data.pdbj.org/pub/pdb/validation_reports/n9/5n97 | HTTPS FTP |
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-Related structure data
Related structure data | 3604MC 5n8pC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
#1: Protein | Mass: 73159.305 Da / Num. of mol.: 6 / Fragment: UNP residues 249-1026 / Source method: isolated from a natural source / Source: (natural) Caulobacter crescentus NA1000 (bacteria) / Variant: YB2811 / References: UniProt: A0A0H3C8J1 #2: Chemical | ChemComp-CA / |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: subtomogram averaging |
Crystal symmetry | ∠γ: 1 ° / C sampling length: 10 Å / A: 220 Å / B: 220 Å / C: 220 Å / Space group name H-M: P121 |
-Sample preparation
Component | Name: Caulobacter crescentus S-layer / Type: ORGANELLE OR CELLULAR COMPONENT / Details: Caulobacter crescentus S-layer / Entity ID: #1 / Source: NATURAL |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Caulobacter crescentus NA1000 (bacteria) / Strain: YB2811 / Cellular location: extra-cellular |
Buffer solution | pH: 7 / Details: PYE medium |
Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Caulobacter crescentus stalk |
Specimen support | Grid material: COPPER/RHODIUM / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R2/2 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 283.15 K / Details: 1.5 s blot |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 105000 X / Calibrated magnification: 105000 X / Nominal defocus max: 5000 nm / Nominal defocus min: 2000 nm / Calibrated defocus min: 2000 nm / Calibrated defocus max: 5000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 70 K / Temperature (min): 70 K |
Image recording | Average exposure time: 1 sec. / Electron dose: 3.4 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Details: Dose symmetric tilt scheme (Hagen et al) |
EM imaging optics | Energyfilter name: GIF Quantum / Energyfilter upper: 20 eV / Energyfilter lower: 0 eV |
-Processing
EM software |
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Crystal symmetry | ∠γ: 1 ° / C sampling length: 10 Å / A: 220 Å / B: 220 Å / C: 220 Å / Space group name H-M: P121 | ||||||||||||||||
CTF correction | Details: Following Schur et al, 2016 / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||
3D reconstruction | Resolution: 7.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 51866 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: 2D CRYSTAL | ||||||||||||||||
EM volume selection | Method: RELIONList of Walmart brands / Details: RELION subtomogram averaging / Num. of tomograms: 110 / Num. of volumes extracted: 51866 / Reference model: Ab initio | ||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL / Target criteria: Cross-correlation coefficient |