The DOI matches its Crossref record; 1 of the 1 references it cites were found in a registry or an index.
Detection of Structural Dynamics by FRET: A Photon Distribution and Fluorescence Lifetime Analysis of Systems with Multiple States
Abstract
Two complementary methods in confocal single-molecule fluorescence spectroscopy are presented to analyze conformational dynamics by Forster resonance energy transfer (FRET) measurements considering simulated and experimental data. First, an extension of photon distribution analysis (PDA) is applied to characterize conformational exchange between two or more states via global analysis of the shape of FRET peaks for different time bins. PDA accurately predicts the shape of FRET efficiency histograms in the presence of FRET fluctuations, taking into account shot noise and background contributions. Dynamic-PDA quantitatively recovers FRET efficiencies of the interconverting states and relaxation times of dynamics on the time scale of the diffusion time t(d) (typically milliseconds), with a dynamic range of the method of about +/-1 order of magnitude with respect to t(d). Correction procedures are proposed to consider the factors limiting the accuracy of dynamic-PDA, such as brightness variations, shortening of the observation time due to diffusion, and a contribution of multimolecular events. Second, an analysis procedure for multiparameter fluorescence detection is presented, where intensity-derived FRET efficiency is correlated with the fluorescence lifetime of the donor quenched by FRET. If a maximum likelihood estimator is applied to compute a mean fluorescence lifetime of mixed states, one obtains a fluorescence weighted mean lifetime. Thus a mixed state is detected by a characteristic shift of the fluorescence lifetime, which becomes longer than that expected for a single species with the same intensity-derived FRET efficiency. Analysis tools for direct visual inspection of two-dimensional diagrams of FRET efficiency versus donor lifetime are presented for the cases of static and dynamic FRET. Finally these new techniques are compared with fluorescence correlation spectroscopy.
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Margittai, M., Widengren, J., Schweinberger, E., Schröder, G. F., Felekyan, S., Haustein, E., König, M., Fasshauer, D., Grubmüller, H., Jahn, R., et al. (2003) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.2331232100 · Cited by 290
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Cited by
Precision and accuracy of single-molecule FRET measurements—a multi-laboratory benchmark study
Hellenkamp, B., Schmid, S., Doroshenko, O., Opanasyuk, O., Kühnemuth, R., Rezaei Adariani, S., Ambrose, B., Aznauryan, M., Barth, A., Birkedal, V., et al. (2018) · Nature Methods · doi:10.1038/s41592-018-0085-0 · Cited by 519
Self-citationAccurate Single-Molecule FRET Studies Using Multiparameter Fluorescence Detection
Sisamakis, E., Valeri, A., Kalinin, S., Rothwell, P. J., & Seidel, C. A. M. (2010) · Methods in Enzymology · doi:10.1016/s0076-6879(10)75018-7 · Cited by 268
Self-citationSindbert, S., Kalinin, S., Nguyen, H., Kienzler, A., Clima, L., Bannwarth, W., Appel, B., Müller, S., & Seidel, C. A. M. (2011) · Journal of the American Chemical Society · doi:10.1021/ja105725e · Cited by 268
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Metadata
- Authors
- Kalinin, S., Valeri, A., Antonik, M., Felekyan, S., & Seidel, C. A. M.
- Venue
- The Journal of Physical Chemistry B
- Publisher
- American Chemical Society (ACS)
- Published
- 2010-05-20
- Volume
- 114
- Issue
- 23
- Pages
- 7983-7995
- ISSN
- 1520-5207 (ISSN-L)
- 1520-6106
- OpenAlex
- W2073341329
- Cited by
- 195
- References
- 1
- FWCI
- 7.28
- Type
- journal article
- Language
- en
- Open access
- closed
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