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Higuera-Rodriguez, R.A.* ; De Pascali, M.C.* ; Aziz, M. ; Sattler, M. ; Rant, U.* ; Kaiser, W.J.*

Kinetic FRET assay to measure binding-induced conformational changes of nucleic acids.

ACS sens. 8, 4597-4606 (2023)
Verlagsversion DOI PMC
Open Access Gold (Paid Option)
Creative Commons Lizenzvertrag
The interaction of small molecules or proteins with RNA or DNA often involves changes in the nucleic acid (NA) folding and structure. A biophysical characterization of these processes helps us to understand the underlying molecular mechanisms. Here, we propose kinFRET (kinetics Förster resonance energy transfer), a real-time ensemble FRET methodology to measure binding and folding kinetics. With kinFRET, the kinetics of conformational changes of NAs (DNA or RNA) upon analyte binding can be directly followed via a FRET signal using a chip-based biosensor. We demonstrate the utility of this approach with two representative examples. First, we monitored the conformational changes of different formats of an aptamer (MN19) upon interaction with small-molecule analytes. Second, we characterized the binding kinetics of RNA recognition by tandem K homology (KH) domains of the human insulin-like growth factor II mRNA-binding protein 3 (IMP3), which reveals distinct kinetic contributions of the two KH domains. Our data demonstrate that kinFRET is well suited to study the kinetics and conformational changes of NA-analyte interactions.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Korrespondenzautor
Schlagwörter Förster Resonance Energy Transfer ; Rna Binding Protein ; Aptamer ; Binding Kinetics ; Biosensors ; Nucleic Acid Conformational Changes ; Switchsense Technology; Stem Length; Rna; Cocaine; Protein; Recognition; Dynamics; Affinity; Aptamer.; Nmr
ISSN (print) / ISBN 2379-3694
e-ISSN 2379-3694
Zeitschrift ACS sensors
Quellenangaben Band: 8, Heft: 12, Seiten: 4597-4606 Artikelnummer: , Supplement: ,
Verlag ACS
Verlagsort Washington, DC
Nichtpatentliteratur Publikationen
Begutachtungsstatus Peer reviewed
Institut(e) Institute of Structural Biology (STB)
Förderungen German Research Foundation (DFG)
European Unions Horizon 2020 research and innovation program