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bioRxiv · 10.1101/2022.05.23.493026

Identification and Quantification of Within-Burst Dynamics in Singly-Labeled Single-Molecule Fluorescence Lifetime Experiments

Harris, P. D.·Lerner, E.
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Abstract

Single-molecule spectroscopy has revolutionized molecular biophysics and provided means to probe how structural moieties within biomolecules spatially reorganize at different timescales. There are several single-molecule methodologies that probe local structural dynamics in the vicinity of a single dye-labeled residue, which rely on fluorescence lifetimes as readout. Nevertheless, an analytical framework to quantify dynamics in such single-molecule single-dye fluorescence bursts, at timescales of microseconds to milliseconds, has not yet been demonstrated. Here, we suggest an analytical framework for identifying and quantifying within-burst lifetime-based dynamics, such as conformational dynamics recorded in single-molecule photo-isomerization related fluorescence enhancement. After testing the capabilities of the analysis on simulations, we proceed to exhibit within-burst millisecond local structural dynamics in the unbound -synuclein monomer. The analytical framework provided in this work paves the way for extracting a full picture of the energy landscape for the coordinate probed by fluorescence-lifetime based single-molecule measurements.

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Harris, P. D., Lerner, E.. 2022-05-24. Identification and Quantification of Within-Burst Dynamics in Singly-Labeled Single-Molecule Fluorescence Lifetime Experiments. https://doi.org/10.1101/2022.05.23.493026

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