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- PDB-7acy: H/L (SLPH/SLPL) complex from C. difficile (CD630 strain) -

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Basic information

Entry
Database: PDB / ID: 7acy
TitleH/L (SLPH/SLPL) complex from C. difficile (CD630 strain)
Components(S-layer protein) x 2
KeywordsSTRUCTURAL PROTEIN / Bacterial surface / S-layer
Function / homologyLow molecular weight S layer protein, N-terminal / Low molecular weight S layer protein, N-terminal, subdomain / Low molecular weight S layer protein N terminal / Putative cell wall binding repeat 2 / Cell wall binding domain 2 (CWB2) / outer membrane-bounded periplasmic space / identical protein binding / Precursor of the S-layer proteins
Function and homology information
Biological speciesClostridioides difficile (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.55 Å
AuthorsLanzoni-Mangutchi, P. / Barwinska-Sendra, A. / Salgado, P.S.
Funding support United Kingdom, 1items
OrganizationGrant numberCountry
Wellcome Trust204877/Z/16/Z United Kingdom
CitationJournal: Nat Commun / Year: 2022
Title: Structure and assembly of the S-layer in C. difficile.
Authors: Paola Lanzoni-Mangutchi / Oishik Banerji / Jason Wilson / Anna Barwinska-Sendra / Joseph A Kirk / Filipa Vaz / Shauna O'Beirne / Arnaud Baslé / Kamel El Omari / Armin Wagner / Neil F ...Authors: Paola Lanzoni-Mangutchi / Oishik Banerji / Jason Wilson / Anna Barwinska-Sendra / Joseph A Kirk / Filipa Vaz / Shauna O'Beirne / Arnaud Baslé / Kamel El Omari / Armin Wagner / Neil F Fairweather / Gillian R Douce / Per A Bullough / Robert P Fagan / Paula S Salgado /
Abstract: Many bacteria and archaea possess a two-dimensional protein array, or S-layer, that covers the cell surface and plays crucial roles in cell physiology. Here, we report the crystal structure of SlpA, ...Many bacteria and archaea possess a two-dimensional protein array, or S-layer, that covers the cell surface and plays crucial roles in cell physiology. Here, we report the crystal structure of SlpA, the main S-layer protein of the bacterial pathogen Clostridioides difficile, and use electron microscopy to study S-layer organisation and assembly. The SlpA crystal lattice mimics S-layer assembly in the cell, through tiling of triangular prisms above the cell wall, interlocked by distinct ridges facing the environment. Strikingly, the array is very compact, with pores of only ~10 Å in diameter, compared to other S-layers (30-100 Å). The surface-exposed flexible ridges are partially dispensable for overall structure and assembly, although a mutant lacking this region becomes susceptible to lysozyme, an important molecule in host defence. Thus, our work gives insights into S-layer organisation and provides a basis for development of C. difficile-specific therapeutics.
History
DepositionSep 11, 2020Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 9, 2022Provider: repository / Type: Initial release
Revision 1.1Jan 31, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: S-layer protein
B: S-layer protein
C: S-layer protein
D: S-layer protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)147,1787
Polymers146,8904
Non-polymers2883
Water1,74797
1


  • Idetical with deposited unit
  • defined by software
  • Evidence: gel filtration, ELISA assay probed SLPL (LMW SLP) and SLPH (HMW SLP) interaction and confirmed H/L complex formation (Fagan, Mol. Micro, 2009)
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area13070 Å2
ΔGint-106 kcal/mol
Surface area60130 Å2
MethodPISA
Unit cell
Length a, b, c (Å)72.670, 78.250, 81.580
Angle α, β, γ (deg.)81.89, 66.98, 65.26
Int Tables number1
Space group name H-MP1

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Components

#1: Protein S-layer protein /


Mass: 33949.785 Da / Num. of mol.: 2 / Source method: isolated from a natural source
Source: (natural) Clostridioides difficile (strain 630) (bacteria)
Plasmid details: RT012, SLCT7 / Strain: 630 / References: UniProt: Q183M8
#2: Protein S-layer protein /


Mass: 39495.035 Da / Num. of mol.: 2 / Source method: isolated from a natural source
Source: (natural) Clostridioides difficile (strain 630) (bacteria)
Plasmid details: RT012, SLCT7 / Strain: 630 / References: UniProt: Q183M8
#3: Chemical ChemComp-SO4 / SULFATE ION / Sulfate


Mass: 96.063 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: SO4
#4: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 97 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.63 Å3/Da / Density % sol: 53.2 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 6.2
Details: 0.1 M MES pH 6.2 1.25 Lithium chloride 16% PEG 6000 10% Glycerol

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: Diamond / Beamline: I24 / Wavelength: 0.9686 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Sep 28, 2018
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9686 Å / Relative weight: 1
ReflectionResolution: 2.55→52.31 Å / Num. obs: 47230 / % possible obs: 96.4 % / Redundancy: 3.3 % / Biso Wilson estimate: 60.17 Å2 / CC1/2: 0.992 / CC star: 0.998 / Rmerge(I) obs: 0.128 / Rpim(I) all: 0.083 / Rrim(I) all: 0.153 / Net I/σ(I): 7
Reflection shellResolution: 2.55→2.64 Å / Redundancy: 3.4 % / Rmerge(I) obs: 0.704 / Mean I/σ(I) obs: 1.5 / Num. unique obs: 4750 / CC1/2: 0.732 / CC star: 0.919 / Rpim(I) all: 0.445 / Rrim(I) all: 0.836 / % possible all: 97.6

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Processing

Software
NameVersionClassification
REFMAC5.8.0258refinement
DIALSdata reduction
Aimlessdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 3CVZ, 5j6Q, D_129211090/992/994

3cvz

5j6q


Resolution: 2.55→52.31 Å / Cor.coef. Fo:Fc: 0.941 / Cor.coef. Fo:Fc free: 0.924 / SU B: 38.843 / SU ML: 0.353 / Cross valid method: THROUGHOUT / ESU R: 1.11 / ESU R Free: 0.332 / Stereochemistry target values: MAXIMUM LIKELIHOOD
Details: U VALUES : WITH TLS ADDED HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS U VALUES : RESIDUAL ONLY
RfactorNum. reflection% reflectionSelection details
Rfree0.26 2432 5.2 %RANDOM
Rwork0.23 ---
obs0.23 44767 96.4 %-
Solvent computationIon probe radii: 0.7 Å / Shrinkage radii: 0.7 Å / VDW probe radii: 1.1 Å / Solvent model: MASK
Displacement parametersBiso mean: 60.166 Å2
Baniso -1Baniso -2Baniso -3
1--1.1 Å20.68 Å2-0.19 Å2
2--1.05 Å20.54 Å2
3----0.13 Å2
Refinement stepCycle: LAST / Resolution: 2.55→52.31 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms10306 0 15 97 10418
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0040.01910416
X-RAY DIFFRACTIONr_bond_other_d0.0010.029890
X-RAY DIFFRACTIONr_angle_refined_deg0.9161.90914092
X-RAY DIFFRACTIONr_angle_other_deg0.9172.93223094
X-RAY DIFFRACTIONr_dihedral_angle_1_deg4.69951378
X-RAY DIFFRACTIONr_dihedral_angle_2_deg34.64527.286420
X-RAY DIFFRACTIONr_dihedral_angle_3_deg12.727151900
X-RAY DIFFRACTIONr_dihedral_angle_4_deg18.4641522
X-RAY DIFFRACTIONr_chiral_restr0.050.21696
X-RAY DIFFRACTIONr_gen_planes_refined0.0020.0211644
X-RAY DIFFRACTIONr_gen_planes_other0.0010.021746
X-RAY DIFFRACTIONr_mcbond_it1.0053.035524
X-RAY DIFFRACTIONr_mcbond_other1.0053.035523
X-RAY DIFFRACTIONr_mcangle_it1.7884.5426898
X-RAY DIFFRACTIONr_mcangle_other1.7884.5436899
X-RAY DIFFRACTIONr_scbond_it0.8083.1114892
X-RAY DIFFRACTIONr_scbond_other0.7963.0884880
X-RAY DIFFRACTIONr_scangle_other1.4154.5937176
X-RAY DIFFRACTIONr_long_range_B_refined3.53136.34411150
X-RAY DIFFRACTIONr_long_range_B_other3.5336.33311146
LS refinement shellResolution: 2.55→2.616 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.413 200 -
Rwork0.373 3349 -
obs--97.58 %
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
10.40610.0542-0.35740.2171-0.33391.0013-0.019-0.036-0.14220.1457-0.03290.0144-0.1894-0.23330.0520.652-0.1232-0.22970.32770.08440.1794-7.686-2.13247.01
21.29780.79290.36041.8616-0.3330.880.06160.12430.11170.0276-0.0433-0.07780.06870.0597-0.01840.5007-0.1571-0.19680.07630.09140.122233.39939.52424.957
30.6101-0.26160.33280.4376-0.28560.455-0.0912-0.1360.0979-0.07570.04420.0108-0.0546-0.32780.04690.5518-0.1041-0.24980.43080.04080.11885.63783.128-11.359
40.25630.3425-0.0582.0162-0.03170.5152-0.08150.0194-0.01840.03280.07690.0765-0.05440.07590.00460.4447-0.2093-0.20280.11270.10970.12799.99321.138-8.99
Refinement TLS group
IDRefine-IDRefine TLS-IDAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1A1 - 318
2X-RAY DIFFRACTION2B2 - 374
3X-RAY DIFFRACTION3C1 - 318
4X-RAY DIFFRACTION4D2 - 374

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