Model-free photon analysis of diffusion-based single-molecule FRET experiments

Abstract Photon-by-photon analysis tools for diffusion-based single-molecule Förster resonance energy transfer (smFRET) experiments often describe protein dynamics with Markov models. However, FRET efficiencies are only projections of the conformational space such that the measured dynamics can appe...

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Main Authors: Ivan Terterov, Daniel Nettels, Tanya Lastiza-Male, Kim Bartels, Christian Löw, Renee Vancraenenbroeck, Itay Carmel, Gabriel Rosenblum, Hagen Hofmann
Format: Article
Language:English
Published: Nature Portfolio 2025-07-01
Series:Nature Communications
Online Access:https://doi.org/10.1038/s41467-025-60764-8
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author Ivan Terterov
Daniel Nettels
Tanya Lastiza-Male
Kim Bartels
Christian Löw
Renee Vancraenenbroeck
Itay Carmel
Gabriel Rosenblum
Hagen Hofmann
author_facet Ivan Terterov
Daniel Nettels
Tanya Lastiza-Male
Kim Bartels
Christian Löw
Renee Vancraenenbroeck
Itay Carmel
Gabriel Rosenblum
Hagen Hofmann
author_sort Ivan Terterov
collection DOAJ
description Abstract Photon-by-photon analysis tools for diffusion-based single-molecule Förster resonance energy transfer (smFRET) experiments often describe protein dynamics with Markov models. However, FRET efficiencies are only projections of the conformational space such that the measured dynamics can appear non-Markovian. Model-free methods to quantify FRET efficiency fluctuations would be desirable in this case. Here, we present such an approach. We determine FRET efficiency correlation functions free of artifacts from the finite length of photon trajectories or the diffusion of molecules through the confocal volume. We show that these functions capture the dynamics of proteins from nano- to milliseconds both in simulation and experiment, which provides a rigorous validation of current model-based analysis approaches.
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spelling doaj-art-5fdbdb46fb5246108073f033d75f7b152025-08-20T04:01:40ZengNature PortfolioNature Communications2041-17232025-07-0116111610.1038/s41467-025-60764-8Model-free photon analysis of diffusion-based single-molecule FRET experimentsIvan Terterov0Daniel Nettels1Tanya Lastiza-Male2Kim Bartels3Christian Löw4Renee Vancraenenbroeck5Itay Carmel6Gabriel Rosenblum7Hagen Hofmann8Department of Chemical and Structural Biology, Weizmann Institute of ScienceDepartment of Biochemistry, University of ZurichDepartment of Chemical and Structural Biology, Weizmann Institute of ScienceCentre for Structural Systems Biology (CSSB) DESYCentre for Structural Systems Biology (CSSB) DESYDepartment of Chemical and Structural Biology, Weizmann Institute of ScienceDepartment of Chemical and Structural Biology, Weizmann Institute of ScienceDepartment of Chemical and Structural Biology, Weizmann Institute of ScienceDepartment of Chemical and Structural Biology, Weizmann Institute of ScienceAbstract Photon-by-photon analysis tools for diffusion-based single-molecule Förster resonance energy transfer (smFRET) experiments often describe protein dynamics with Markov models. However, FRET efficiencies are only projections of the conformational space such that the measured dynamics can appear non-Markovian. Model-free methods to quantify FRET efficiency fluctuations would be desirable in this case. Here, we present such an approach. We determine FRET efficiency correlation functions free of artifacts from the finite length of photon trajectories or the diffusion of molecules through the confocal volume. We show that these functions capture the dynamics of proteins from nano- to milliseconds both in simulation and experiment, which provides a rigorous validation of current model-based analysis approaches.https://doi.org/10.1038/s41467-025-60764-8
spellingShingle Ivan Terterov
Daniel Nettels
Tanya Lastiza-Male
Kim Bartels
Christian Löw
Renee Vancraenenbroeck
Itay Carmel
Gabriel Rosenblum
Hagen Hofmann
Model-free photon analysis of diffusion-based single-molecule FRET experiments
Nature Communications
title Model-free photon analysis of diffusion-based single-molecule FRET experiments
title_full Model-free photon analysis of diffusion-based single-molecule FRET experiments
title_fullStr Model-free photon analysis of diffusion-based single-molecule FRET experiments
title_full_unstemmed Model-free photon analysis of diffusion-based single-molecule FRET experiments
title_short Model-free photon analysis of diffusion-based single-molecule FRET experiments
title_sort model free photon analysis of diffusion based single molecule fret experiments
url https://doi.org/10.1038/s41467-025-60764-8
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