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TitleTarget ssDNA activates the NADase activity of prokaryotic SPARTA immune system.
Journal, issue, pagesNat Chem Biol, Vol. 20, Issue 4, Page 503-511, Year 2024
Publish dateNov 6, 2023
AuthorsJun-Tao Zhang / Xin-Yang Wei / Ning Cui / Ruilin Tian / Ning Jia /
PubMed AbstractArgonaute proteins (Agos), which use small RNAs or DNAs as guides to recognize complementary nucleic acid targets, mediate RNA silencing in eukaryotes. In prokaryotes, Agos are involved in immunity: ...Argonaute proteins (Agos), which use small RNAs or DNAs as guides to recognize complementary nucleic acid targets, mediate RNA silencing in eukaryotes. In prokaryotes, Agos are involved in immunity: the short prokaryotic Ago/TIR-APAZ (SPARTA) immune system triggers cell death by degrading NAD in response to invading plasmids, but its molecular mechanisms remain unknown. Here we used cryo-electron microscopy to determine the structures of inactive monomeric and active tetrameric Crenotalea thermophila SPARTA complexes, revealing mechanisms underlying SPARTA assembly, RNA-guided recognition of target single-stranded DNA (ssDNA) and subsequent SPARTA tetramerization, as well as tetramerization-dependent NADase activation. The small RNA guides Ago to recognize its ssDNA target, inducing SPARTA tetramerization via both Ago- and TIR-mediated interactions and resulting in a two-stranded, parallel, head-to-tail TIR rearrangement primed for NAD hydrolysis. Our findings thus identify the molecular basis for target ssDNA-mediated SPARTA activation, which will facilitate the development of SPARTA-based biotechnological tools.
External linksNat Chem Biol / PubMed:37932528
MethodsEM (single particle)
Resolution2.54 - 3.11 Å
Structure data

35419

8ifk

EMDB-35419, PDB-8ifk:
Cryo-EM structure of monomeric SPARTA gRNA-ssDNA target complex
Method: EM (single particle) / Resolution: 2.54 Å

35420

8ifl

EMDB-35420: Cryo-EM structure of tetrameric SPARTA gRNA-ssDNA target complex in state1
PDB-8ifl: Cryo-EM structure of tetrameric SPARTA gRNA-ssDNA target complex in state 1
Method: EM (single particle) / Resolution: 3.11 Å

35421

8ifm

EMDB-35421, PDB-8ifm:
Cryo-EM structure of tetrameric SPARTA gRNA-ssDNA target complex in state 2
Method: EM (single particle) / Resolution: 2.92 Å

36843

8k34

EMDB-36843, PDB-8k34:
Cryo-EM structure of SPARTA gRNA binary complex
Method: EM (single particle) / Resolution: 2.81 Å

Chemicals

ChemComp-MG:
Unknown entry

Source
  • thermoflavifilum thermophilum (bacteria)
  • escherichia coli 'bl21-gold(de3)plyss ag' (bacteria)
KeywordsRNA BINDING PROTEIN/RNA/DNA / RNA BINDING PROTEIN-RNA-DNA COMPLEX / STRUCTURAL PROTEIN/RNA / STRUCTURAL PROTEIN-RNA COMPLEX

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