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- PDB-8ykf: The DSR2-DSAD1 complex with DSAD1 on the opposite sides -

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Basic information

Entry
Database: PDB / ID: 8ykf
TitleThe DSR2-DSAD1 complex with DSAD1 on the opposite sides
Components
  • DSAD1
  • SIR2-like domain-containing protein
KeywordsIMMUNE SYSTEM / Complex
Function / homologySIR2-like domain / SIR2-like domain / DHS-like NAD/FAD-binding domain superfamily / symbiont-mediated suppression of host innate immune response / SIR2-like domain-containing protein / DSR anti-defense 1
Function and homology information
Biological speciesBacillus subtilis (bacteria)
Bacillus phage SPBc2 (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.35 Å
AuthorsZheng, J. / Yang, X.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32070040, 32370071 China
CitationJournal: Int J Biol Macromol / Year: 2024
Title: Structural insights into autoinhibition and activation of defense-associated sirtuin protein.
Authors: Xu Yang / Yiqun Wang / Jianting Zheng /
Abstract: Bacterial defense-associated sirtuin 2 (DSR2) proteins harbor an N-terminal sirtuin (SIR2) domain degrading NAD. DSR2 from Bacillus subtilis 29R is autoinhibited and unable to hydrolyze NAD in the ...Bacterial defense-associated sirtuin 2 (DSR2) proteins harbor an N-terminal sirtuin (SIR2) domain degrading NAD. DSR2 from Bacillus subtilis 29R is autoinhibited and unable to hydrolyze NAD in the absence of phage infection. A tail tube protein (TTP) of phage SPR activates the DSR2 while a DSR2-inhibiting protein of phage SPbeta, known as DSAD1 (DSR anti-defense 1), inactivates the DSR2. Although DSR2 structures in complexed with TTP and DSAD1, respectively, have been reported recently, the autoinhibition and activation mechanisms remain incompletely understood. Here, we present cryo-electron microscopy structures of the DSR2-NAD complex in autoinhibited state and the in vitro assembled DSR2-TFD (TTP tube-forming domain) complex in activated state. The DSR2-NAD complex reveals that the autoinhibited DSR2 assembles into an inactive tetramer, binding NAD through a distinct pocket situated outside active site. Binding of TFD into cavities within the sensor domains of DSR2 triggers a conformational change in SIR2 regions, activating its NADase activity, whereas the TTP β-sandwich domain (BSD) is flexible and does not contribute to the activation process. The activated form of DSR2 exists as tetramers and dimers, with the tetramers exhibiting more NADase activity. Overall, our results extend the current understanding of autoinhibition and activation of DSR2 immune proteins.
History
DepositionMar 5, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Aug 14, 2024Provider: repository / Type: Initial release
Revision 1.1Oct 9, 2024Group: Data collection / Database references ...Data collection / Database references / Source and taxonomy / Structure summary
Category: em_admin / entity_src_gen ...em_admin / entity_src_gen / pdbx_entry_details / struct_ref / struct_ref_seq / struct_ref_seq_dif
Item: _em_admin.last_update / _entity_src_gen.gene_src_common_name ..._em_admin.last_update / _entity_src_gen.gene_src_common_name / _entity_src_gen.pdbx_gene_src_gene / _entity_src_gen.pdbx_gene_src_scientific_name / _struct_ref.db_code / _struct_ref.pdbx_db_accession / _struct_ref.pdbx_seq_one_letter_code / _struct_ref_seq.pdbx_db_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: SIR2-like domain-containing protein
B: SIR2-like domain-containing protein
C: SIR2-like domain-containing protein
D: SIR2-like domain-containing protein
E: DSAD1
F: DSAD1


Theoretical massNumber of molelcules
Total (without water)502,2896
Polymers502,2896
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
SIR2-like domain-containing protein


Mass: 118635.789 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus subtilis (bacteria) / Gene: BSNT_07056 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: D4G637
#2: Protein DSAD1


Mass: 13872.930 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus phage SPBc2 (virus) / Gene: yotI / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: O64191
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: The DSR2-DSAD1 complex with DSAD1 on the opposite sides
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
11Bacillus subtilis (bacteria)1423
21Bacillus phage SPbeta (virus)2932878
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMTris bufferTris-HCl1
2150 mMSodium chlorideNaCl1
32 mMDithiothreitolDTT1
42 %GlycerolGlycerol1
SpecimenConc.: 2.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 289 K

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Electron microscopy imaging

MicroscopyModel: FEI TITAN
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 40 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

EM softwareName: EPU / Category: image acquisition
CTF correctionType: NONE
3D reconstructionResolution: 3.35 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 72614 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00735822
ELECTRON MICROSCOPYf_angle_d0.66948262
ELECTRON MICROSCOPYf_dihedral_angle_d4.2754632
ELECTRON MICROSCOPYf_chiral_restr0.0385126
ELECTRON MICROSCOPYf_plane_restr0.0036160

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