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- PDB-8ps6: Crystal structure of the N-terminal domain of SduA -

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Basic information

Entry
Database: PDB / ID: 8ps6
TitleCrystal structure of the N-terminal domain of SduA
ComponentsShedu effector protein
KeywordsIMMUNE SYSTEM / SduA / Shedu / Prokaryotic immune system
Biological speciesEscherichia coli KTE10 (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.52 Å
AuthorsLoeff, L. / Walter, A. / Jinek, M.
Funding supportEuropean Union, 2items
OrganizationGrant numberCountry
European Research Council (ERC)820152European Union
H2020 Marie Curie Actions of the European Commission845268European Union
CitationJournal: Cell / Year: 2025
Title: DNA end sensing and cleavage by the Shedu anti-phage defense system.
Authors: Luuk Loeff / Alexander Walter / Gian Tizio Rosalen / Martin Jinek /
Abstract: The detection of molecular patterns associated with invading pathogens is a hallmark of innate immune systems. Prokaryotes deploy sophisticated host defense mechanisms in innate anti-phage immunity. ...The detection of molecular patterns associated with invading pathogens is a hallmark of innate immune systems. Prokaryotes deploy sophisticated host defense mechanisms in innate anti-phage immunity. Shedu is a single-component defense system comprising a putative nuclease SduA. Here, we report cryoelectron microscopy (cryo-EM) structures of apo- and double-stranded DNA (dsDNA)-bound tetrameric SduA assemblies, revealing that the N-terminal domains of SduA form a clamp that recognizes free DNA ends. End binding positions the DNA over the PD-(D/E)XK nuclease domain, resulting in dsDNA nicking at a fixed distance from the 5' end. The end-directed DNA nicking activity of Shedu prevents propagation of linear DNA in vivo. Finally, we show that phages escape Shedu immunity by suppressing their recombination-dependent DNA replication pathway. Taken together, these results define the antiviral mechanism of Shedu systems, underlining the paradigm that recognition of pathogen-specific nucleic acid structures is a conserved feature of innate immunity across all domains of life.
History
DepositionJul 13, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jan 22, 2025Provider: repository / Type: Initial release
Revision 1.1Feb 19, 2025Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Shedu effector protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)22,2633
Polymers22,0711
Non-polymers1922
Water59433
1
A: Shedu effector protein
hetero molecules

A: Shedu effector protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)44,5266
Polymers44,1422
Non-polymers3844
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation6_556-x,-x+y,-z+11
Buried area3810 Å2
ΔGint-77 kcal/mol
Surface area21110 Å2
MethodPISA
Unit cell
Length a, b, c (Å)225.543, 225.543, 32.029
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number155
Space group name H-MH32
Space group name HallR32"
Symmetry operation#1: x,y,z
#2: -y,x-y,z
#3: -x+y,-x,z
#4: x-y,-y,-z
#5: -x,-x+y,-z
#6: y,x,-z
#7: x+1/3,y+2/3,z+2/3
#8: -y+1/3,x-y+2/3,z+2/3
#9: -x+y+1/3,-x+2/3,z+2/3
#10: x-y+1/3,-y+2/3,-z+2/3
#11: -x+1/3,-x+y+2/3,-z+2/3
#12: y+1/3,x+2/3,-z+2/3
#13: x+2/3,y+1/3,z+1/3
#14: -y+2/3,x-y+1/3,z+1/3
#15: -x+y+2/3,-x+1/3,z+1/3
#16: x-y+2/3,-y+1/3,-z+1/3
#17: -x+2/3,-x+y+1/3,-z+1/3
#18: y+2/3,x+1/3,-z+1/3
Components on special symmetry positions
IDModelComponents
11A-332-

HOH

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Components

#1: Protein Shedu effector protein / Shedu protein / DUF4263 domain-containing protein


Mass: 22070.945 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli KTE10 (bacteria) / Gene: SduA / Production host: Escherichia coli BL21(DE3) (bacteria)
#2: Chemical ChemComp-SO4 / SULFATE ION


Mass: 96.063 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: SO4
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 33 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestN
Has protein modificationN

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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: 3.7 Å3/Da / Density % sol: 66.79 %
Crystal growTemperature: 277.15 K / Method: vapor diffusion, hanging drop / pH: 8
Details: HEPES 100 mM, Ammonium Sulfate 550 mM, protein concentration: 4.5 mg /ml, 1 uL reservoir + 1 uL protein solution.

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SLS / Beamline: X06DA / Wavelength: 0.99999 Å
DetectorType: DECTRIS PILATUS 2M-F / Detector: PIXEL / Date: Aug 21, 2021
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.99999 Å / Relative weight: 1
ReflectionResolution: 2.518→112.772 Å / Num. obs: 10123 / % possible obs: 95.8 % / Redundancy: 20.3 % / Biso Wilson estimate: 49.15 Å2 / Rmerge(I) obs: 0.174 / Rpim(I) all: 0.039 / Rrim(I) all: 0.178 / Net I/σ(I): 14
Reflection shellResolution: 2.518→2.561 Å / Redundancy: 21.5 % / Num. unique obs: 518 / CC1/2: 0.748 / % possible all: 100

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Processing

Software
NameVersionClassification
PHENIX1.20.1_4487refinement
Aimlessdata scaling
PHASERphasing
XDSdata reduction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.52→42.62 Å / SU ML: 0.3953 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 31.1651
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflection
Rfree0.2445 461 4.56 %
Rwork0.2224 9656 -
obs0.2235 10117 95.77 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 59.81 Å2
Refinement stepCycle: LAST / Resolution: 2.52→42.62 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1485 0 10 33 1528
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00871526
X-RAY DIFFRACTIONf_angle_d1.19512064
X-RAY DIFFRACTIONf_chiral_restr0.0911222
X-RAY DIFFRACTIONf_plane_restr0.0045267
X-RAY DIFFRACTIONf_dihedral_angle_d13.9547566
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.52-2.560.33581350.28093222X-RAY DIFFRACTION96
2.88-3.630.27881710.25963222X-RAY DIFFRACTION97.03
3.63-42.620.21161550.19073212X-RAY DIFFRACTION94.31
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)
14.65927755323-1.47477695843-2.713602576584.399059441511.006367961038.6645042451-0.2891471683430.0218675515062-0.08296452058040.256036377948-0.03489127971070.483774565380.177846863002-0.8125881882560.3387936323610.495613324931-0.03243966271140.003925642909760.370158983276-0.07647345661210.315223877215-12.294889444256.709658725116.2828883918
24.549218939360.384918466493-1.078406230298.356140556710.397544827625.12885118892-0.2608627317320.187974326233-0.34248484508-0.314635210319-0.0837634425982-0.2445519923460.2198251998650.08067841505660.3204807170390.3644124035150.07352066068030.1071768992460.2803264815970.01032506434070.304005368086-0.32042253933341.52435064317.90992399754
33.800473992184.65746687501-2.761728093123.95210314015-4.447794113924.2292760505-0.3336257574730.21205371681-0.451886255882-0.6805718524560.287791225647-0.4869415258610.604811635645-0.5102913634140.0808695677710.5720290289660.05803709347860.08850245254540.4229396936130.1090357035520.665712173884-26.2580340689.1121764996322.0782235116
Refinement TLS group

Refine-ID: X-RAY DIFFRACTION / Auth asym-ID: A / Label asym-ID: A

IDRefine TLS-IDSelection detailsAuth seq-IDLabel seq-ID
11chain 'A' and (resid 1 through 72 )1 - 721 - 68
22chain 'A' and (resid 73 through 148 )73 - 14869 - 144
33chain 'A' and (resid 149 through 185 )149 - 184145 - 180

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