title: Structural analysis of 5'-mRNA-cap interactions with the human AGO2 MID domain.
abstract: In RNA silencing, microRNA (miRNA)-mediated translational repression occurs through mechanisms that do not invoke messenger-RNA (mRNA) target cleavage by Argonaute proteins. The nature of these mechanisms is unclear, but several recent studies have proposed that a direct interaction between the mRNA-cap and the middle (MID) domain of Argonautes is involved. Here, we present crystallographic and NMR data demonstrating that cap analogues do not bind significantly to the isolated MID domain of human Argonaute 2 (hAGO2) and are found in the miRNA 5'-nucleotide binding site in an implausible binding mode. Additionally, in vitro pull-down experiments with full-length hAGO2 indicate that the interaction with cap analogues is nonspecific.
authors.affiliation: Department of Biochemistry, McGill University, 1160 Pine Avenue West, Montreal, Quebec H3A 1A3, Canada.
authors.forename: Filipp
authors.initials: F
authors.lastname: Frank
authors.forename: Marc R
authors.initials: MR
authors.lastname: Fabian
authors.forename: Janusz
authors.initials: J
authors.lastname: Stepinski
authors.forename: Jacek
authors.initials: J
authors.lastname: Jemielity
authors.forename: Edward
authors.initials: E
authors.lastname: Darzynkiewicz
authors.forename: Nahum
authors.initials: N
authors.lastname: Sonenberg
authors.forename: Bhushan
authors.initials: B
authors.lastname: Nagar
mesh_terms: D060565:Argonaute Proteins; D001703:Biophysics; D003461:Crystallography; D015852:Eukaryotic Initiation Factor-2 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary / Q000235:genetics*; D012315:RNA Caps / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference
chemical_list: C466422:AGO2 protein, human; D060565:Argonaute Proteins; D015852:Eukaryotic Initiation Factor-2; D012315:RNA Caps