Single Pair Förster Resonance Energy Transfer: A Versatile Tool To Investigate Protein Conformational Dynamics |
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Authors: | Lena Voith von Voithenberg Don C. Lamb |
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Affiliation: | 1. Department Chemie, Center for Nanoscience (CeNS), Center for Integrated Protein Science Munich (CIPSM), Nanosystem Initiative Munich (NIM), Ludwig‐Maximilians‐Universit?t München, Butenandtstr. 5–13, 81377 München, Germany;2. BIOSS Centre for Signalling Studies, Sch?nzlestr. 18, 79104 Freiburg, Germany |
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Abstract: | Conformational changes of proteins and other biomolecules play a fundamental role in their functional mechanism. Single pair Förster resonance energy transfer (spFRET) offers the possibility to detect these conformational changes and dynamics, and to characterize their underlying kinetics. Using spFRET on microscopes with different modes of detection, dynamic timescales ranging from nanoseconds to seconds can be quantified. Confocal microscopy can be used as a means to analyze dynamics in the range of nanoseconds to milliseconds, while total internal reflection fluorescence (TIRF) microscopy offers information about conformational changes on timescales of milliseconds to seconds. While the existence of dynamics can be directly inferred from the FRET efficiency time trace or the correlation of FRET efficiency and fluorescence lifetime, additional computational approaches are required to extract the kinetic rates of these dynamics, a short overview of which is given in this review. |
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Keywords: | conformational dynamics FCS fluorescence spectroscopy FRET single molecule TIRF |
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