A blind benchmark of analysis tools to infer kinetic rate constants from single-molecule FRET trajectories

Journal article (2022)

Authors

Markus Götz CNRS/Université de Montpellier II
A. Barth Heinrich Heine University
Søren S.R. Bohr Faculty of Health and Medical Sciences, University of Copenhagen
Richard Börner University of Zürich
Jixin Chen Ohio University
Thorben Cordes Ludwig Maximilians University
C.V. de Lannoy Wageningen University & Research
Dick de Ridder Wageningen University & Research
Sonja Schmid Wageningen University & Research
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Published Date

2022

Language

English

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Abstract

Single-molecule FRET (smFRET) is a versatile technique to study the dynamics and function of biomolecules since it makes nanoscale movements detectable as fluorescence signals. The powerful ability to infer quantitative kinetic information from smFRET data is, however, complicated by experimental limitations. Diverse analysis tools have been developed to overcome these hurdles but a systematic comparison is lacking. Here, we report the results of a blind benchmark study assessing eleven analysis tools used to infer kinetic rate constants from smFRET trajectories. We test them against simulated and experimental data containing the most prominent difficulties encountered in analyzing smFRET experiments: different noise levels, varied model complexity, non-equilibrium dynamics, and kinetic heterogeneity. Our results highlight the current strengths and limitations in inferring kinetic information from smFRET trajectories. In addition, we formulate concrete recommendations and identify key targets for future developments, aimed to advance our understanding of biomolecular dynamics through quantitative experiment-derived models.