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Nature. 2009 Oct 8;461(7265):754-61. doi: 10.1038/nature08434.

Nucleation, propagation and cleavage of target RNAs in Ago silencing complexes.

Author information

1
Structural Biology Program, Memorial-Sloan Kettering Cancer Center, New York, New York 10065, USA.

Abstract

The slicer activity of the RNA-induced silencing complex resides within its Argonaute (Ago) component, in which the PIWI domain provides the catalytic residues governing guide-strand mediated site-specific cleavage of target RNA. Here we report on structures of ternary complexes of Thermus thermophilus Ago catalytic mutants with 5'-phosphorylated 21-nucleotide guide DNA and complementary target RNAs of 12, 15 and 19 nucleotides in length, which define the molecular basis for Mg(2+)-facilitated site-specific cleavage of the target. We observe pivot-like domain movements within the Ago scaffold on proceeding from nucleation to propagation steps of guide-target duplex formation, with duplex zippering beyond one turn of the helix requiring the release of the 3'-end of the guide from the PAZ pocket. Cleavage assays on targets of various lengths supported this model, and sugar-phosphate-backbone-modified target strands showed the importance of structural and catalytic divalent metal ions observed in the crystal structures.

PMID:
19812667
PMCID:
PMC2880917
DOI:
10.1038/nature08434
[Indexed for MEDLINE]
Free PMC Article

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