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- EMDB-3604: Structure of the C. crescentus S-layer -

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Basic information

Entry
Database: EMDB / ID: 3604
TitleStructure of the C. crescentus S-layer
Map dataSub-tomogram averaging of the Caulobacter crescentus S-layer
SampleCaulobacter crescentus S-layer
  • S-layer protein rsaA
  • ligand
Function / homologyHaemolysin-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
SourceCaulobacter crescentus na1000 / / bacteria
Methodsubtomogram averaging / cryo EM / 7.4 Å resolution
AuthorsBharat TA / Hagen WJ
CitationJournal: 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 ReportPDB-ID: 5n97

SummaryFull reportAbout validation report
DateDeposition: Feb 24, 2017 / Header (metadata) release: Mar 29, 2017 / Map release: Apr 19, 2017 / Last update: Aug 2, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.11
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.11
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: : PDB-5n97
  • Surface level: 0.11
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_3604.map.gz (map file in CCP4 format, 32001 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
200 pix
1.35 Å/pix.
= 270. Å
200 pix
1.35 Å/pix.
= 270. Å
200 pix
1.35 Å/pix.
= 270. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.35 Å
Density
Contour Level:0.11 (by author), 0.11 (movie #1):
Minimum - Maximum-0.25166476 - 0.5049828
Average (Standard dev.)-0.009878825 (0.0565414)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions200200200
Origin000
Limit199199199
Spacing200200200
CellA=B=C: 270 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.351.351.35
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z270.000270.000270.000
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.2520.505-0.010

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Supplemental data

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Sample components

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Entire Caulobacter crescentus S-layer

EntireName: Caulobacter crescentus S-layer / Details: Caulobacter crescentus S-layer / Number of components: 3

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Component #1: cellular-component, Caulobacter crescentus S-layer

Cellular-componentName: Caulobacter crescentus S-layer / Details: Caulobacter crescentus S-layer / Recombinant expression: No
SourceSpecies: Caulobacter crescentus na1000 / / bacteria / Strain: YB2811
Source (natural)Location in cell: extra-cellular

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Component #2: protein, S-layer protein rsaA

ProteinName: S-layer protein rsaA / Recombinant expression: No
MassTheoretical: 73.159305 kDa
SourceSpecies: Caulobacter crescentus na1000 / / bacteria

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Component #3: ligand, CALCIUM ION

LigandName: CALCIUM IONCalcium / Number of Copies: 114 / Recombinant expression: No
MassTheoretical: 4.007805 MDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionSpecimen conc.: 1 mg/ml / Buffer solution: PYE medium / pH: 7
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 283.15 K / Humidity: 100 % / Details: 1.5 s blot

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 3.4 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 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 HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER / Tilt Angle: -60 - 60 deg. / Temperature: K ( 70 - 70 K)
CameraDetector: GATAN K2 (4k x 4k)

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Image acquisition

Image acquisitionDetails: Dose symmetric tilt scheme (Hagen et al)

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Image processing

ProcessingMethod: subtomogram averaging
3D reconstructionAlgorithm: 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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Atomic model buiding

Output model

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