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Crystal structure and ligand binding of the MID domain of a eukaryotic Argonaute protein

Department of Biochemistry, Max Planck Institute for Developmental Biology, Spemannstrasse 35, Tübingen D-72076, Germany.
EMBO Reports (Impact Factor: 7.86). 07/2010; 11(7):522-7. DOI: 10.1038/embor.2010.81
Source: PubMed

ABSTRACT Argonaute (AGO) proteins are core components of RNA-induced silencing complexes and have essential roles in RNA-mediated gene silencing. They are characterized by a bilobal architecture, consisting of one lobe containing the amino-terminal and PAZ domains and another containing the MID and PIWI domains. Except for the PAZ domain, structural information on eukaryotic AGO domains is not yet available. In this study, we report the crystal structure of the MID domain of the eukaryotic AGO protein QDE-2 from Neurospora crassa. This domain adopts a Rossmann-like fold and recognizes the 5'-terminal nucleotide of a guide RNA in a manner similar to its prokaryotic counterparts. The 5'-nucleotide-binding site shares common residues with a second, adjacent ligand-binding site, suggesting a mechanism for the cooperative binding of ligands to the MID domain of eukaryotic AGOs.

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Available from: Andreas Boland, Aug 21, 2014
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    • "Except for the 5 0 -phosphate binding pocket for siRNA or miRNA, Djuranovic et al. [2010] proposed that Dm_AGO1 has a second miRNA-dependent site that can bind nucleotides such as the 5 0 -cap. The second ligand-binding site of Dm_AGO1 might be under allosteric control, whereas the other AGO proteins might be regulated by distinct ligand or might no longer be regulated by a second ligand [Boland et al., 2010]. According to this, it induces us to speculate that the second sulfate ion in Nc_QDE2 MID domain may occupy the second ligand binding site. "
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    ABSTRACT: Argonaute (AGO) proteins are highly specialized small-RNA-binding modules and small RNAs are anchored to their specific binding pockets guiding AGO proteins to target mRNA molecules for silencing or destruction. The 135 full-length AGO protein sequences derived from 36 species covering prokaryote, archaea, and eukaryote are chosen for structural and functional analyses. The results show that bacteria and archaeal AGO proteins are clustered in the same clade and there exist multiple AGO proteins in most eukaryotic species, demonstrating that the increase of AGO gene copy number and horizontal gene transfer (HGT) have been the main evolutionary driving forces for adaptability and biodiversity. And the emergence of PAZ domain in AGO proteins is the unique evolutionary event. The analysis of middle domain (MID)-nucleotide contaction shows that either the position of sulfate I bond in Nc_QDE2 or the site of phosphate I bond in Hs_AGO2 represents the 5'-nucleotide binding site of miRNA. Also, H334, T335, and Y336 of Hs_AGO1 can form hydrogen bonds with 3'-overhanging ends of miRNAs and the same situation exists in Hs_AGO2, Hs_AGO3, Hs_AGO4, Dm_AGO1, and Ce_Alg1. Some PIWI domains containing conserved DDH motif have no slicer activity, and post-translational modifications may be associated with the endonucleolytic activities of AGOs. With the numbers of AGO genes increasing and fewer crystal structures available, the evolutionary and functional analyses of AGO proteins can help clarify the molecular mechanism of function diversification in response to environmental changes, and solve major issues including host defense mechanism against virus infection and molecular basis of disease.
    Journal of Cellular Biochemistry 08/2012; 113(8):2576-85. DOI:10.1002/jcb.24133 · 3.37 Impact Factor
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    • "In contrast to the conserved conformations of residues directly contacting the 5 0 -phosphate, the nucleotide specificity loops in AtAGO1, AtAGO2 and AtAGO5 all have distinct, rigid conformations (Figure 2D; Supplementary Table 4; Supplementary Figure 8), which is in accordance with the lack of conservation observed in their sequences (Supplementary Figure 2). A comparison of this loop in all available structures of eukaryotic MID domains, including the AtAGO structures determined here, hAGO2 and Neurospora crassa AGO (QDE-2) (Boland et al, 2010), demonstrates that this loop can assume variable conformations between species (Supplementary Figure 9). Fittingly, with the exception of QDE-2 for which no data are available on 5 0 -bias, each of these AGOs displays distinct 5 0 -nucleotide biases in their associated small RNAs. "
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    ABSTRACT: The 5'-nucleotide of small RNAs associates directly with the MID domain of Argonaute (AGO) proteins. In humans, the identity of the 5'-base is sensed by the MID domain nucleotide specificity loop and regulates the integrity of miRNAs. In Arabidopsis thaliana, the 5'-nucleotide also controls sorting of small RNAs into the appropriate member of the AGO family; however, the structural basis for this mechanism is unknown. Here, we present crystal structures of the MID domain from three Arabidopsis AGOs, AtAGO1, AtAGO2 and AtAGO5, and characterize their interactions with nucleoside monophosphates (NMPs). In AtAGOs, the nucleotide specificity loop also senses the identity of the 5'-nucleotide but uses more diverse modes of recognition owing to the greater complexity of small RNAs found in plants. Binding analyses of these interactions reveal a strong correlation between their affinities and evolutionary conservation.
    The EMBO Journal 07/2012; 31(17):3588-95. DOI:10.1038/emboj.2012.204 · 10.75 Impact Factor
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    • "In this study, the bioinformatics was used to analyze the domain spatial conformation of Schistosoma Argonaute protein, and this laid the initial foundation for further study on the function of Argonaute protein in the growth and development of Schistosoma. According to relevant literature [12] [13] [14] [15] , the Schistosoma Argonaute protein is a highly conserved family member, and it is a multi-domain protein which contains the N-terminal domain, PAZ domain, MID domain and Piwi domain. Through the analysis in this study, the functional expression form comprised the N-terminal PAZ domain and C-terminal Piwi domain for the Argonaute protein domain's spatial conformation was obtained. "
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    ABSTRACT: To analyze the amino acid sequence composition, secondary structure, the spatial conformation of its domain and other characteristics of Argonaute protein. Bioinformatics tools and the internet server were used. Firstly, the amino acid sequence composition features of the Argonaute protein were analyzed, and the phylogenetic tree was constructed. Secondly, Argonaute protein's distribution of secondary structure and its physicochemical properties were predicted. Lastly, the protein functional expression form of the domain group was established through the Phyre-based analysis on the spatial conformation of Argonaute protein domains. 593 amino acids were encoded by Argonaute protein, the phylogenetic tree was constructed, and Argonaute protein's distribution of secondary structure and its physicochemical properties were obtained through analysis. In addition, the functional expression form which comprised the N-terminal PAZ domain and C-terminal Piwi domain for the Argonaute protein was obtained with Phyre. The information relationship between the structure and function of the Argonaute protein can be initially established with bioinformatics tools and the internet server, and this provides the theoretical basis for further clarifying the function of Schistosoma Argonaute protein.
    Asian Pacific Journal of Tropical Biomedicine 08/2011; 1(4):275-8. DOI:10.1016/S2221-1691(11)60042-7
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