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- PDB-7acz: RdeltaD2 H/L (LMW SLP/HMW SLP) complex from C. difficile SlpA (R2... -

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Basic information

Entry
Database: PDB / ID: 7acz
TitleRdeltaD2 H/L (LMW SLP/HMW SLP) complex from C. difficile SlpA (R20291 strain)
Components
  • SLPH (HMW SLP)
  • SLPL deltaD2 (LMW SLP D2 truncation),SLPL deltaD2 (LMW SLP D2 truncation)
KeywordsSTRUCTURAL PROTEIN / Bacterial surface / S-layer
Function / homologyLow molecular weight S-layer protein, domain 1 / Dna Ligase; domain 1 / 2-Layer Sandwich / Alpha Beta / :
Function and homology information
Biological speciesClostridioides difficile (bacteria)
Clostridioides difficile R20291 (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.5 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 1.1Jun 19, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: SLPL deltaD2 (LMW SLP D2 truncation),SLPL deltaD2 (LMW SLP D2 truncation)
B: SLPH (HMW SLP)
C: SLPL deltaD2 (LMW SLP D2 truncation),SLPL deltaD2 (LMW SLP D2 truncation)
D: SLPH (HMW SLP)


Theoretical massNumber of molelcules
Total (without water)126,6954
Polymers126,6954
Non-polymers00
Water00
1
A: SLPL deltaD2 (LMW SLP D2 truncation),SLPL deltaD2 (LMW SLP D2 truncation)
B: SLPH (HMW SLP)


Theoretical massNumber of molelcules
Total (without water)63,3482
Polymers63,3482
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4900 Å2
ΔGint-44 kcal/mol
Surface area24010 Å2
MethodPISA
2
C: SLPL deltaD2 (LMW SLP D2 truncation),SLPL deltaD2 (LMW SLP D2 truncation)
D: SLPH (HMW SLP)


Theoretical massNumber of molelcules
Total (without water)63,3482
Polymers63,3482
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4210 Å2
ΔGint-27 kcal/mol
Surface area24130 Å2
MethodPISA
Unit cell
Length a, b, c (Å)52.745, 80.352, 81.920
Angle α, β, γ (deg.)97.030, 90.219, 90.217
Int Tables number1
Space group name H-MP1
Space group name HallP1
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1(chain "A" and (resid 5 or (resid 6 and (name...
d_2ens_1(chain "C" and (resid 5 through 60 or (resid 61...
d_1ens_2(chain "B" and ((resid 3 through 4 and (name N...
d_2ens_2(chain "D" and (resid 3 through 19 or (resid 20...

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg label comp-IDEnd label comp-IDLabel asym-IDLabel seq-ID
d_11ens_1SERVALA1 - 50
d_12ens_1SERSERA55 - 86
d_13ens_1ILEGLYA88 - 131
d_14ens_1VALTHRA134 - 153
d_21ens_1SERTHRC1 - 146
d_11ens_2LYSASPB1 - 181
d_12ens_2LYSGLYB183 - 344
d_13ens_2ASNMETB346 - 405
d_21ens_2LYSGLYD1 - 343
d_22ens_2ASNMETD346 - 405

NCS ensembles :
ID
ens_1
ens_2

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Components

#1: Protein SLPL deltaD2 (LMW SLP D2 truncation),SLPL deltaD2 (LMW SLP D2 truncation) / Cell surface protein (S-layer protein)


Mass: 19262.543 Da / Num. of mol.: 2
Fragment: SLPL deltaD2 (LMW SLP D2 truncation) from C. difficile (R20291),SLPL deltaD2 (LMW SLP D2 truncation)
Mutation: deleted aa 115-259
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Clostridioides difficile (strain R20291) (bacteria)
Strain: R20291 / Gene: slpA, CDR20291_2682 / Plasmid details: RT027, SLCT4 / Production host: Clostridioides difficile R20291 (bacteria) / References: UniProt: C9YQ17
#2: Protein SLPH (HMW SLP) / Cell surface protein (S-layer protein)


Mass: 44085.027 Da / Num. of mol.: 2 / Fragment: SLPH (HMW SLP) from C. difficile (R20291) / Source method: isolated from a natural source
Source: (natural) Clostridioides difficile R20291 (bacteria)
Plasmid details: RT027, SLCT4 / References: UniProt: C9YQ17
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: 2.72 Å3/Da / Density % sol: 54.87 %
Crystal growTemperature: 293.15 K / Method: vapor diffusion, sitting drop / pH: 6
Details: 0.1 M MES pH 6.0 0.875 M Lithium chloride 11.5% PEG 6,000 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: 3.5→52.74 Å / Num. obs: 16691 / % possible obs: 98.75 % / Redundancy: 3.4 % / Biso Wilson estimate: 37.95 Å2
Details: The Rmerge is very high for this crystal due to merging of data from 2 highly anisotropic crystals, that are also not completely isomorphous. However, this was necessary to generate ...Details: The Rmerge is very high for this crystal due to merging of data from 2 highly anisotropic crystals, that are also not completely isomorphous. However, this was necessary to generate interpretable electron density maps, as data from either crystal separately was enough.
CC1/2: 0.616 / CC star: 0.873 / Rmerge(I) obs: 0.5102 / Rpim(I) all: 0.3238 / Rrim(I) all: 0.6068 / Net I/σ(I): 10
Reflection shellResolution: 3.5→3.63 Å / Redundancy: 3.5 % / Rmerge(I) obs: 0.9043 / Mean I/σ(I) obs: 2.47 / Num. unique obs: 1620 / CC1/2: 0.438 / CC star: 0.78 / Rpim(I) all: 0.5631 / Rrim(I) all: 1.069 / % possible all: 97.03

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Processing

Software
NameVersionClassification
PHENIX1.18.2_3874refinement
PHENIX1.18.2_3874refinement
DIALSdata reduction
Aimlessdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: Dep D_19292110995

Resolution: 3.5→52.74 Å / SU ML: 0.4221 / Cross valid method: FREE R-VALUE / σ(F): 1.96 / Phase error: 31.17
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflectionSelection details
Rfree0.2786 1528 5.18 %RANDOM
Rwork0.2544 27992 --
obs0.2557 16606 87.78 %-
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 37 Å2
Refinement stepCycle: LAST / Resolution: 3.5→52.74 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms7890 0 0 0 7890
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00257951
X-RAY DIFFRACTIONf_angle_d0.595110798
X-RAY DIFFRACTIONf_chiral_restr0.04291344
X-RAY DIFFRACTIONf_plane_restr0.00261408
X-RAY DIFFRACTIONf_dihedral_angle_d6.71691149
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDRefine-IDTypeRms dev position (Å)
ens_1d_2AX-RAY DIFFRACTIONTorsion NCS0.815686536469
ens_2d_2BX-RAY DIFFRACTIONTorsion NCS0.703614709263
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.5-3.610.28861620.2982449X-RAY DIFFRACTION86.92
3.61-3.740.4261190.28942544X-RAY DIFFRACTION85.41
3.74-3.890.37421760.29122479X-RAY DIFFRACTION87.22
3.89-4.070.25041440.26982550X-RAY DIFFRACTION88.21
4.07-4.280.29261680.24462546X-RAY DIFFRACTION89.16
4.28-4.550.19781110.23412565X-RAY DIFFRACTION87.39
4.55-4.90.31151490.23142531X-RAY DIFFRACTION87.52
4.9-5.40.2461370.23742601X-RAY DIFFRACTION89.07
5.4-6.170.2488860.2712594X-RAY DIFFRACTION88.45
6.18-7.770.2633960.2712617X-RAY DIFFRACTION88.6
7.78-52.740.22911800.21162516X-RAY DIFFRACTION87.7
Refinement TLS params.Method: refined / Origin x: 6.83834376163 Å / Origin y: -36.3952231107 Å / Origin z: 10.2700128284 Å
111213212223313233
T0.0390504095105 Å20.000404378431586 Å20.0291962208907 Å2-0.138962305232 Å20.0103175473504 Å2--0.142071510976 Å2
L-0.00467041492349 °2-0.0222730303971 °20.0669154606771 °2-0.596893231454 °20.0814241540752 °2--0.652473365991 °2
S0.0399590873126 Å °0.00351802425101 Å °-0.0251390437869 Å °-0.0137774976384 Å °-0.00638624473251 Å °-0.0114293875421 Å °-0.00589432448504 Å °-0.0226061021223 Å °0.000655995992817 Å °
Refinement TLS groupSelection details: all

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