Abstract
Förster resonance energy transfer (FRET) is a versatile tool to study the conformational dynamics of proteins. Here, we describe the use of confocal and total internal reflection fluorescence (TIRF) microscopy to follow the conformational cycling of DEAD-box helicases on the single molecule level, using the eukaryotic translation initiation factor eIF4A as an illustrative example. Confocal microscopy enables the study of donor-acceptor-labeled molecules in solution, revealing the population of different conformational states present. With TIRF microscopy, surface-immobilized molecules can be imaged as a function of time, revealing sequences of conformational states and the kinetics of conformational changes.
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Krause, L., Klostermeier, D. (2021). Probing RNA Helicase Conformational Changes by Single-Molecule FRET Microscopy. In: Boudvillain, M. (eds) RNA Remodeling Proteins. Methods in Molecular Biology, vol 2209. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0935-4_8
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DOI: https://doi.org/10.1007/978-1-0716-0935-4_8
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