1z26
Structure of Pyrococcus furiosus Argonaute with bound tungstate
Structural highlights
FunctionAGO_PYRFU A DNA-guided ssDNA endonuclease that may play a role in defense against invading mobile genetic elements. Uses short 5'-phospho-ssDNA sequences as guides (gDNA) to bind complementary target strands, resulting in cleavage of the target DNA (tDNA). Endonucleolytically cleaves DNA in short dsDNA (the gDNA indicates where to cleave on the tDNA). Efficient guide-dependent target DNA cleavage requires a minimal gDNA length of 15 nucleotides (nt) and works up to at least 31 nt. Overexpression decreases plasmid transformation efficiency. Has no appreciable activity with gRNA or on target RNA. Also has guide-independent activity on plasmid DNA called 'chopping' (PubMed:25925567). The cleavage site is 10 nucleotides (nt) downstream of the target residue base-paired with the 5'-end of the gDNA, cleavage is insensitive to adenine methylation. DNA cleavage produces 5'-phosphomonoesters (as it can be ligated by T4 DNA ligase) (PubMed:28165224).[1] [2] Evolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedGenetic, biochemical and structural studies have implicated Argonaute proteins as the catalytic core of the RNAi effector complex, RISC. Here we show that recombinant, human Argonaute2 can combine with a small interfering RNA (siRNA) to form minimal RISC that accurately cleaves substrate RNAs. Recombinant RISC shows many of the properties of RISC purified from human or Drosophila melanogaster cells but also has surprising features. It shows no stimulation by ATP, suggesting that factors promoting product release are missing from the recombinant enzyme. The active site is made up of a unique Asp-Asp-His (DDH) motif. In the RISC reconstitution system, the siRNA 5' phosphate is important for the stability and the fidelity of the complex but is not essential for the creation of an active enzyme. These studies demonstrate that Argonaute proteins catalyze mRNA cleavage within RISC and provide a source of recombinant enzyme for detailed biochemical studies of the RNAi effector complex. Purified Argonaute2 and an siRNA form recombinant human RISC.,Rivas FV, Tolia NH, Song JJ, Aragon JP, Liu J, Hannon GJ, Joshua-Tor L Nat Struct Mol Biol. 2005 Apr;12(4):340-9. Epub 2005 Mar 30. PMID:15800637[3] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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