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- EMDB-65406: Helical structure of gRNA-tDNA SPARDA complex -

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Basic information

Entry
Database: EMDB / ID: EMD-65406
TitleHelical structure of gRNA-tDNA SPARDA complex
Map dataThe main Map of SPARDA's Role in Filament Formation
Sample
  • Complex: gRNA-tDNA SPARDA complex
    • DNA: target DNA
    • RNA: guide RNA
    • Protein or peptide: pAGO
    • Protein or peptide: DREN-APAZ
    • DNA: substrate ssDNA
    • RNA: substrate ssRNA
KeywordsCryo-EM / SPARDA / pAgo / CELL INVASION
Biological speciesNovosphingopyxis baekryungensis (bacteria) / Bacteriophage sp. (virus)
Methodhelical reconstruction / cryo EM / Resolution: 2.64 Å
AuthorsLi Y / Zheng Q / Li S / Jiang Y
Funding support China, 1 items
OrganizationGrant numberCountry
Not funded China
CitationJournal: Nat Commun / Year: 2026
Title: Filament assembly induced by the recognition of target DNA activates the prokaryotic Argonaute SPARDA system.
Authors: Wanyue Zhang / Yuchen Jiang / Yu Li / Xiangkai Zhen / Shuying Xu / Ning-Shao Xia / Shaowei Li / Xurong Wang / Qingbing Zheng / Songying Ouyang /
Abstract: The short prokaryotic Argonaute (pAgo) proteins, in conjunction with their associated effector molecules, constitute a defence mechanism in prokaryotes that protects against phage infections. The ...The short prokaryotic Argonaute (pAgo) proteins, in conjunction with their associated effector molecules, constitute a defence mechanism in prokaryotes that protects against phage infections. The SPARDA is characterized by its collateral nuclease activity, which, upon activation through target DNA recognition, non-specifically cleaves a wide range of nucleic acid substrates. Nevertheless, the structural underpinnings of its collateral activity have remained elusive. In this study, we investigate the NbaSPARDA system from Novosphingopyxis baekryungensis and reveal that RNA-guided DNA recognition triggers the assembly of a higher-order filamentous structure. This filamentation is essential for the tetramerization of the DREN nuclease domain, which in turn facilitates the accumulation and cleavage of substrate nucleic acids. Through the determination of the gRNA-bound and RNA-DNA duplex-bound cryo-EM structures, we delineate a sequential monomer-dimer-monomer-filament transition during SPARDA activation. These insights collectively elucidate a filament-dependent activation mechanism underpinning the short pAgo-mediated immune response, which is crucial for antiviral defence.
History
DepositionJul 18, 2025-
Header (metadata) releaseJan 14, 2026-
Map releaseJan 14, 2026-
UpdateJan 21, 2026-
Current statusJan 21, 2026Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_65406.png

Downloads & links

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Map

FileDownload / File: emd_65406.map.gz / Format: CCP4 / Size: 1.4 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThe main Map of SPARDA's Role in Filament Formation
Voxel sizeX=Y=Z: 0.65 Å
Density
Contour LevelBy AUTHOR: 0.095
Minimum - Maximum-0.4171389 - 0.9718239
Average (Standard dev.)0.0011496993 (±0.021954887)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions720720720
Spacing720720720
CellA=B=C: 467.99997 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: The half Map of SPARDA's Role in Filament Formation

Fileemd_65406_half_map_1.map
AnnotationThe half Map of SPARDA's Role in Filament Formation
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: The half Map of SPARDA's Role in Filament Formation

Fileemd_65406_half_map_2.map
AnnotationThe half Map of SPARDA's Role in Filament Formation
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : gRNA-tDNA SPARDA complex

EntireName: gRNA-tDNA SPARDA complex
Components
  • Complex: gRNA-tDNA SPARDA complex
    • DNA: target DNA
    • RNA: guide RNA
    • Protein or peptide: pAGO
    • Protein or peptide: DREN-APAZ
    • DNA: substrate ssDNA
    • RNA: substrate ssRNA

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Supramolecule #1: gRNA-tDNA SPARDA complex

SupramoleculeName: gRNA-tDNA SPARDA complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#6
Source (natural)Organism: Novosphingopyxis baekryungensis (bacteria)

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Macromolecule #1: target DNA

MacromoleculeName: target DNA / type: dna / ID: 1 / Number of copies: 4 / Classification: DNA
Source (natural)Organism: Bacteriophage sp. (virus)
Molecular weightTheoretical: 6.139976 KDa
SequenceString:
(DT)(DA)(DT)(DC)(DG)(DT)(DC)(DA)(DG)(DC) (DT)(DG)(DT)(DG)(DC)(DA)(DG)(DT)(DA)(DT)

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Macromolecule #5: substrate ssDNA

MacromoleculeName: substrate ssDNA / type: dna / ID: 5 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Bacteriophage sp. (virus)
Molecular weightTheoretical: 3.364208 KDa
SequenceString:
(DC)(DT)(DG)(DT)(DG)(DC)(DA)(DG)(DT)(DA) (DT)

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Macromolecule #2: guide RNA

MacromoleculeName: guide RNA / type: rna / ID: 2 / Number of copies: 4
Source (natural)Organism: Novosphingopyxis baekryungensis (bacteria)
Molecular weightTheoretical: 6.390879 KDa
SequenceString:
AUACUGCACA GCUGACGAUA

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Macromolecule #6: substrate ssRNA

MacromoleculeName: substrate ssRNA / type: rna / ID: 6 / Number of copies: 1
Source (natural)Organism: Novosphingopyxis baekryungensis (bacteria)
Molecular weightTheoretical: 3.46613 KDa
SequenceString:
UACUGCACAG C

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Macromolecule #3: pAGO

MacromoleculeName: pAGO / type: protein_or_peptide / ID: 3 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Novosphingopyxis baekryungensis (bacteria)
Molecular weightTheoretical: 54.492809 KDa
Recombinant expressionOrganism: Novosphingopyxis baekryungensis (bacteria)
SequenceString: MTFETRIFDE PELEFGDHHH HQDPRLGLSE AGPLQTFLGD VIKIGVVGNS KTIEDTRKFI ETVSSGVEGK GEKHPNMHPP FPGLGNQSP YRCRFEIEDG ATAALTKSKL DKIGKEPDHY RAVEMAVDEI IGELQAMDDG GSRPDVAIIA LPVKLLERVW N AKVDARGT ...String:
MTFETRIFDE PELEFGDHHH HQDPRLGLSE AGPLQTFLGD VIKIGVVGNS KTIEDTRKFI ETVSSGVEGK GEKHPNMHPP FPGLGNQSP YRCRFEIEDG ATAALTKSKL DKIGKEPDHY RAVEMAVDEI IGELQAMDDG GSRPDVAIIA LPVKLLERVW N AKVDARGT TEKSDSSGSD APNFRGMLKA KAMGLSFPIQ IVWEDVIDDK VTIPQKVKES SSRKIQDIAG RTWNLMTSLY YK GSGRIPW RRMPLEGEFS ACYVGISFYR EADGQQLFTS AAQMFDERGR GFVLKGRRAR TESRGRHPYM AREDAKKIIE DVL AAYKLH HKTLPARVFI LKTSRFKDEE ADGIIAALDE AGTELRDLVW VQESYTARIL RDGNYPVLRG TFVDLHGKGL LYTS GSMPY YGTYPGKYDP NPLLLCPHHT SESTVAQLAE EIFSLTKVNW NSTQMNQRLP IPIRAARKVG EVLKYVGEGE VISAD YRKY I

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Macromolecule #4: DREN-APAZ

MacromoleculeName: DREN-APAZ / type: protein_or_peptide / ID: 4 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Novosphingopyxis baekryungensis (bacteria)
Molecular weightTheoretical: 50.419645 KDa
Recombinant expressionOrganism: Bacteria Latreille et al. 1825 (Bacteria stick insect)
SequenceString: MTKKITANQI IGEIGENEVR GRFLTLGWQF DGRSRLEAGI DGIAEVMNEG QPMARMIAVQ IKSTKEGKYT SESDTSFTYL LRTQDLAYW RGSNLPVIVV FYRQSDHSFY WKEVSRDAGP GERRLNIDKV ADLFNASTVN KLAALTVPKT GLGYYVPPLG G GEDALINM ...String:
MTKKITANQI IGEIGENEVR GRFLTLGWQF DGRSRLEAGI DGIAEVMNEG QPMARMIAVQ IKSTKEGKYT SESDTSFTYL LRTQDLAYW RGSNLPVIVV FYRQSDHSFY WKEVSRDAGP GERRLNIDKV ADLFNASTVN KLAALTVPKT GLGYYVPPLG G GEDALINM LPLTLPNEMY IASTTYEPRK AIAVILNGDG PKRFDWVING GTFWSFHDPR TSACSEIVDI DQVEAINTKE LA LHDDIDE QNRFSHLLRQ TLRYQTDSDL GWDKDHKALY FRAIEREVSR NFAYTSSKKK TDANVVSVFK NSKDETRVSF VRH HAFSPR FELMADQWYL IITPTYYYTT NGYAPHQFAA PLLAGKKRLD KSAALRGQVI MWHRFLTQSD HEDLFHSEET PEAY LMFGE PPSIHLDVRV PEDGWVKEKV KRIDEAAQGE GLFSDDI

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statehelical array

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Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 48.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 130.0 Å
Applied symmetry - Helical parameters - Δ&Phi: 80 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 2.64 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 57938
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER
Final angle assignmentType: MAXIMUM LIKELIHOOD

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