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Structure paper

TitleTetramerization-dependent activation of the Sir2-associated short prokaryotic Argonaute immune system.
Journal, issue, pagesNat Commun, Vol. 15, Issue 1, Page 8610, Year 2024
Publish dateOct 4, 2024
AuthorsNing Cui / Jun-Tao Zhang / Zhuolin Li / Xin-Yang Wei / Jie Wang / Ning Jia /
PubMed AbstractEukaryotic Argonaute proteins (eAgos) utilize short nucleic acid guides to target complementary sequences for RNA silencing, while prokaryotic Agos (pAgos) provide immunity against invading plasmids ...Eukaryotic Argonaute proteins (eAgos) utilize short nucleic acid guides to target complementary sequences for RNA silencing, while prokaryotic Agos (pAgos) provide immunity against invading plasmids or bacteriophages. The Sir2-domain associated short pAgo (SPARSA) immune system defends against invaders by depleting NAD and triggering cell death. However, the molecular mechanism underlying SPARSA activation remains unknown. Here, we present cryo-EM structures of inactive monomeric, active tetrameric and active NAD-bound tetrameric SPARSA complexes, elucidating mechanisms underlying SPARSA assembly, guide RNA preference, target ssDNA-triggered SPARSA tetramerization, and tetrameric-dependent NADase activation. Short pAgos form heterodimers with Sir2-APAZ, favoring short guide RNA with a 5'-AU from ColE-like plasmids. RNA-guided recognition of the target ssDNA triggers SPARSA tetramerization via pAgo- and Sir2-mediated interactions. The resulting tetrameric Sir2 rearrangement aligns catalytic residue H186 for NAD hydrolysis. These insights advance our understanding of Sir2-domain associated pAgos immune systems and should facilitate the development of a short pAgo-associated biotechnological toolbox.
External linksNat Commun / PubMed:39366953 / PubMed Central
MethodsEM (single particle)
Resolution2.57 - 3.66 Å
Structure data

39027

8y7z

EMDB-39027, PDB-8y7z:
Cryo-EM structure of the monomeric SPARSA gRNA-ssDNA complex
Method: EM (single particle) / Resolution: 2.57 Å

39028

8y80

EMDB-39028, PDB-8y80:
Cryo-EM structure of the tetrameric SPARSA gRNA-ssDNA complex
Method: EM (single particle) / Resolution: 3.38 Å

39030

8y82

EMDB-39030, PDB-8y82:
Cryo-EM structure of the tetrameric SPARSA gRNA-ssDNA-NAD+ complex
Method: EM (single particle) / Resolution: 3.4 Å

EMDB-39031: Cryo-EM structure of the monomeric SPARSA complex
Method: EM (single particle) / Resolution: 3.66 Å

Chemicals

ChemComp-MG:
Unknown entry

ChemComp-AMP:
ADENOSINE MONOPHOSPHATE / AMP*YM

Source
  • geobacter sulfurreducens (bacteria)
  • escherichia coli 'bl21-gold(de3)plyss ag' (bacteria)
KeywordsSTRUCTURAL PROTEIN/RNA/DNA / RNA BINDING PROTEIN / STRUCTURAL PROTEIN / STRUCTURAL PROTEIN-RNA-DNA complex / RNA BINDING PROTEIN-RNA-DNA COMPLEX

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