Biochemical isolation of Argonaute protein complexes by Ago-APP.

Abstract:

During microRNA (miRNA)-guided gene silencing, Argonaute (Ago) proteins interact with a member of the TNRC6/GW protein family. Here we used a short GW protein-derived peptide fused to GST and demonstrate that it binds to Ago proteins with high affinity. This allows for the simultaneous isolation of all Ago protein complexes expressed in diverse species to identify associated proteins, small RNAs, or target mRNAs. We refer to our method as "Ago protein Affinity Purification by Peptides" (Ago-APP). Furthermore, expression of this peptide competes for endogenous TNRC6 proteins, leading to global inhibition of miRNA function in mammalian cells.

Authors:

J Hauptmann, D Schraivogel, A Bruckmann, S Manickavel, L Jakob, N Eichner, J Pfaff, M Urban, S Sprunck, M Hafner, T Tuschl, R Deutzmann, G Meister

Journal:

Proc Natl Acad Sci U S A

Citation info:

112(38):11841-11845

Publication date:

22nd Sep 2015

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