title: Coupling RNA annealing and strand displacement: a FRET-based microplate reader assay for RNA chaperone activity.
abstract: Proteins with RNA chaperone activity help RNAs to obtain their native conformations, and many of them are active in the two basic reactions-RNA annealing and strand displacement. Therefore, we developed a time-saving in vitro assay that detects protein-facilitated annealing and strand displacement of fluorophore-labeled oligoribonucleotides in a microplate reader The two reactions are followed byfluorescence resonance energy transfer (FRET) in real-time, and the effect of the proteins on the reaction constants can be quantified. The high-throughput property of the fluorescence microplate reader the kinetic characterization, and the material-saving aspect of this assay enables a fast and convenient classification of proteins according to their RNA chaperone activity in annealing and strand displacement.
authors: Max F. Perutz [corrected] Laboratories, University of Vienna, Vienna, Austria.
authors: Lukas
authors: L
authors: Rajkowitsch
authors: Renée
authors: R
authors: Schroeder
mesh_terms: D001665:Binding Sites; D001681:Biological Assay / Q000379:methods; D003199:Computer Systems; D031541:Fluorescence Resonance Energy Transfer / Q000295:instrumentation* / Q000379:methods; D018832:Molecular Chaperones / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D012313:RNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D018832:Molecular Chaperones; D016601:RNA-Binding Proteins; D012313:RNA