|Entry||Database: EMDB / ID: 3604|
|Title||Structure of the C. crescentus S-layer|
|Map data||Sub-tomogram averaging of the Caulobacter crescentus S-layer|
|Sample||Caulobacter crescentus S-layer|
|Function / homology||Haemolysin-type calcium-binding repeat / Serralysin-like metalloprotease, C-terminal / Haemolysin-type calcium-binding repeat (2 copies) / S-layer / cell wall / calcium ion binding / extracellular region / S-layer protein rsaA / S-layer protein|
Function and homology information
|Source||Caulobacter crescentus na1000 / / bacteria|
|Method||subtomogram averaging / cryo EM / 7.4 Å resolution|
|Authors||Bharat TA / Hagen WJ|
|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.
|Validation Report||PDB-ID: 5n97|
SummaryFull reportAbout validation report
|Date||Deposition: Feb 24, 2017 / Header (metadata) release: Mar 29, 2017 / Map release: Apr 19, 2017 / Last update: Aug 2, 2017|
|Structure viewer||EM map: |
Downloads & links
|File||emd_3604.map.gz (map file in CCP4 format, 32001 KB)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 1.35 Å|
CCP4 map header:
-Entire Caulobacter crescentus S-layer
|Entire||Name: Caulobacter crescentus S-layer / Details: Caulobacter crescentus S-layer / Number of components: 3|
-Component #1: cellular-component, Caulobacter crescentus S-layer
|Cellular-component||Name: Caulobacter crescentus S-layer / Details: Caulobacter crescentus S-layer / Recombinant expression: No|
|Source||Species: Caulobacter crescentus na1000 / / bacteria / Strain: YB2811|
|Source (natural)||Location in cell: extra-cellular|
-Component #2: protein, S-layer protein rsaA
|Protein||Name: S-layer protein rsaA / Recombinant expression: No|
|Mass||Theoretical: 73.159305 kDa|
|Source||Species: Caulobacter crescentus na1000 / / bacteria|
-Component #3: ligand, CALCIUM ION
|Ligand||Name: CALCIUM IONCalcium / Number of Copies: 114 / Recombinant expression: No|
|Mass||Theoretical: 4.007805 MDa|
|Specimen||Specimen state: particle / Method: cryo EM|
|Sample solution||Specimen conc.: 1 mg/ml / Buffer solution: PYE medium / pH: 7|
|Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 283.15 K / Humidity: 100 % / Details: 1.5 s blot|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Imaging||Microscope: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 3.4 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Magnification: 105000 X (nominal), 105000 X (calibrated) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 2000 - 5000 nm / Energy filter: GIF Quantum / Energy window: 0-20 eV|
|Specimen Holder||Model: FEI TITAN KRIOS AUTOGRID HOLDER / Tilt Angle: -60 - 60 deg. / Temperature: K ( 70 - 70 K)|
|Camera||Detector: GATAN K2 (4k x 4k)|
|Image acquisition||Details: Dose symmetric tilt scheme (Hagen et al)|
|Processing||Method: subtomogram averaging|
|3D reconstruction||Algorithm: FOURIER SPACE / CTF correction: Following Schur et al, 2016 / Resolution: 7.4 Å / Resolution method: FSC 0.143 CUT-OFF / Euler angles: Following Schur et al, 2016|
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