Structure of yeast Argonaute with guide RNA.

Publication Type:

Journal Article


Nature, Volume 486, Issue 7403, p.368-74 (2012)


Argonaute Proteins, Base Sequence, Biocatalysis, Catalytic Domain, Crystallography, X-Ray, Eukaryotic Cells, Fungal Proteins, Kluyveromyces, Models, Molecular, Molecular Conformation, Molecular Sequence Data, RNA, Guide, Saccharomycetales


<p>The RNA-induced silencing complex, comprising Argonaute and guide RNA, mediates RNA interference. Here we report the 3.2 Å crystal structure of Kluyveromyces polysporus Argonaute (KpAGO) fortuitously complexed with guide RNA originating from small-RNA duplexes autonomously loaded by recombinant KpAGO. Despite their diverse sequences, guide-RNA nucleotides 1-8 are positioned similarly, with sequence-independent contacts to bases, phosphates and 2'-hydroxyl groups pre-organizing the backbone of nucleotides 2-8 in a near-A-form conformation. Compared with prokaryotic Argonautes, KpAGO has numerous surface-exposed insertion segments, with a cluster of conserved insertions repositioning the N domain to enable full propagation of guide-target pairing. Compared with Argonautes in inactive conformations, KpAGO has a hydrogen-bond network that stabilizes an expanded and repositioned loop, which inserts an invariant glutamate into the catalytic pocket. Mutation analyses and analogies to ribonuclease H indicate that insertion of this glutamate finger completes a universally conserved catalytic tetrad, thereby activating Argonaute for RNA cleavage.</p>