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- PDB-9lgw: Cryo-EM structure of SiAgo-Aga1 complex -

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Basic information

Entry
Database: PDB / ID: 9lgw
TitleCryo-EM structure of SiAgo-Aga1 complex
Components
  • Piwi domain-containing protein
  • SiAgo-associated protein1, SiAga1
KeywordsDNA BINDING PROTEIN / Argonaute / RNA BINDING PROTEIN/RNA/DNA / RNA BINDING PROTEIN-RNA-DNA complex
Function / homologyPiwi domain / Piwi domain / Piwi / Ribonuclease H superfamily / nucleic acid binding / Ribonuclease H-like superfamily / Uncharacterized protein / Piwi domain-containing protein
Function and homology information
Biological speciesSaccharolobus islandicus M.16.4 (acidophilic)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsDai, Z.K. / Guan, Z.Y. / Zou, T.T.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Nucleic Acids Res / Year: 2025
Title: Structural and mechanistic insights into the activation of a short prokaryotic argonaute system from archaeon Sulfolobus islandicus.
Authors: Zhikang Dai / Yu Chen / Zeyuan Guan / Xueting Chen / Keyi Tan / Kaiyue Yang / Xuhui Yan / Yidong Liu / Zhou Gong / Wenyuan Han / Tingting Zou /
Abstract: Prokaryotic Argonaute proteins (pAgos) defend the host against invading nucleic acids, including plasmids and viruses. Short pAgo systems confer immunity by inducing cell death upon detecting ...Prokaryotic Argonaute proteins (pAgos) defend the host against invading nucleic acids, including plasmids and viruses. Short pAgo systems confer immunity by inducing cell death upon detecting invading nucleic acids. However, the activation mechanism of the SiAgo system, comprising a short pAgo from the archaeon Sulfolobus islandicus and its associated proteins SiAga1 and SiAga2, remains largely unknown. Here, we determined the cryo-electron microscopy structures of the SiAgo-Aga1 apo complex and the RNA-DNA-bound SiAgo-Aga1 complex at resolutions of 2.7 and 3.0 Å, respectively. Our results revealed that a positively charged pocket is generated from the interaction between SiAgo and SiAga1, exhibiting an architecture similar to APAZ-pAgo of short pAgo systems and accommodating the nucleic acids. Further investigation elucidated the conserved mechanism of nucleic acid recognition by SiAgo-Aga1. Both the SiAgo-Aga1 interaction and nucleic acid recognition by the complex are essential for antiviral defense. Biochemical and structural analyses demonstrated that SiAgo-Aga1 undergoes extensive conformational changes upon binding to the RNA-DNA duplex, thereby licensing its interaction with the effector SiAga2 to trigger the immune response. Overall, our findings highlight the evolutionary conservation of Agos across phylogenetic clades and provide structural insights into the activation mechanism of the SiAgo system.
History
DepositionJan 10, 2025Deposition site: PDBJ / Processing site: PDBC
Revision 1.0Feb 19, 2025Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Piwi domain-containing protein
B: SiAgo-associated protein1, SiAga1


Theoretical massNumber of molelcules
Total (without water)81,8752
Polymers81,8752
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 Piwi domain-containing protein


Mass: 52741.414 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharolobus islandicus M.16.4 (acidophilic)
Gene: M164_1614 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: C4KI01
#2: Protein SiAgo-associated protein1, SiAga1


Mass: 29133.615 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharolobus islandicus M.16.4 (acidophilic)
Gene: M164_1613 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: C4KI00
Has protein modificationY

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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: SiAgo-Aga1 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Saccharolobus islandicus M.16.4 (acidophilic)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1500 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.21.2_5419 / Category: model refinement
CTF correctionType: NONE
3D reconstructionResolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 601826 / Symmetry type: POINT
RefinementHighest resolution: 2.7 Å
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0045055
ELECTRON MICROSCOPYf_angle_d0.5586820
ELECTRON MICROSCOPYf_dihedral_angle_d5.293660
ELECTRON MICROSCOPYf_chiral_restr0.048772
ELECTRON MICROSCOPYf_plane_restr0.004851

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