The crystal structure of human Argonaute2 - PubMed (original) (raw)

The crystal structure of human Argonaute2

Nicole T Schirle et al. Science. 2012.

Abstract

Argonaute proteins form the functional core of the RNA-induced silencing complexes that mediate RNA silencing in eukaryotes. The 2.3 angstrom resolution crystal structure of human Argonaute2 (Ago2) reveals a bilobed molecule with a central cleft for binding guide and target RNAs. Nucleotides 2 to 6 of a heterogeneous mixture of guide RNAs are positioned in an A-form conformation for base pairing with target messenger RNAs. Between nucleotides 6 and 7, there is a kink that may function in microRNA target recognition or release of sliced RNA products. Tandem tryptophan-binding pockets in the PIWI domain define a likely interaction surface for recruitment of glycine-tryptophan-182 (GW182) or other tryptophan-rich cofactors. These results will enable structure-based approaches for harnessing the untapped therapeutic potential of RNA silencing in humans.

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Figures

Figure 1

Figure 1. Structure of human Ago2

A. Schematic of the Ago2 primary sequence. B. Front and top views of Ago2 with the N (purple), PAZ (navy), MID (green), PIWI (grey) domains and linkers L1 (teal) and L2 (blue). A generic guide RNA (red) can be traced for nucleotides 1–8 and 21. Tryptophan molecules (orange) bind to tandem hydrophobic pockets in the PIWI domain.

Figure 2

Figure 2. Comparison of bacterial and human Argonaute structures

A. Superposition of N-PAZ and MID-PIWI lobes of Ago2 (colored as in Fig. 1) onto corresponding lobes from T. thermophiles (yellow). B. Individual domains of Ago2 superimposed on the corresponding domains from T. thermophilus with root-mean-square deviation (rmsd) values for equivalent alpha-carbons indicated. Functional points of interest in Ago2 are labeled.

Figure 3

Figure 3. Conformation of bound guide RNAs

A. The 5′ nucleotides of guide RNAs are recognized by extensive interactions with the MID and PIWI domains. An ordered water molecule is shown as a pink sphere. Hydrogen bonds are shown as dashed orange lines B, C. Ago2 organizes the seed region in an A-form helix by extensive interactions with the phosphate backbone. D. Helix-7 (α7) introduces a kink in the guide RNA between bases 6 and 7 that disrupts helical stacking.

Figure 4

Figure 4. Tandem tryptophan-binding pockets in the PIWI domain

A, B. Residues forming the binding pockets of tryptophan 1 and 2 shown, with hydrogen bonds indicated (dashed, yellow lines). Tryptophan molecules are shown with surrounding unbiased Fo-Fc maps contoured at two sigma (orange). C. Surface representation showing the tryptophan binding pockets. D. Close up view of boxed area in panel C. White dots indicate the shortest direct path connecting the two sites along the surface of Ago2.

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References

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