Search our Database of Scientific Publications and Authors

I’m looking for a
    A flexible approach to assess fluorescence decay functions in complex energy transfer systems.
    BMC Biophys 2015 3;8. Epub 2015 Apr 3.
    Biochemistry and Biology, University of Potsdam, Potsdam, Germany ; Biochemistry and Molecular Biology, University of Hamburg, Hamburg, Germany.
    Background: Time-correlated Förster resonance energy transfer (FRET) probes molecular distances with greater accuracy than intensity-based calculation of FRET efficiency and provides a powerful tool to study biomolecular structure and dynamics. Moreover, time-correlated photon count measurements bear additional information on the variety of donor surroundings allowing more detailed differentiation between distinct structural geometries which are typically inaccessible to general fitting solutions.

    Results: Here we develop a new approach based on Monte Carlo simulations of time-correlated FRET events to estimate the time-correlated single photon counts (TCSPC) histograms in complex systems. This simulation solution assesses the full statistics of time-correlated photon counts and distance distributions of fluorescently labeled biomolecules. The simulations are consistent with the theoretical predictions of the dye behavior in FRET systems with defined dye distances and measurements of randomly distributed dye solutions. We validate the simulation results using a highly heterogeneous aggregation system and explore the conditions to use this tool in complex systems.

    Conclusion: This approach is powerful in distinguishing distance distributions in a wide variety of experimental setups, thus providing a versatile tool to accurately distinguish between different structural assemblies in highly complex systems.

    Similar Publications

    Structural heterogeneity and quantitative FRET efficiency distributions of polyprolines through a hybrid atomistic simulation and Monte Carlo approach.
    PLoS One 2011 24;6(5):e19791. Epub 2011 May 24.
    Theoretical and Computational Biophysics Department, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
    Förster Resonance Energy Transfer (FRET) experiments probe molecular distances via distance dependent energy transfer from an excited donor dye to an acceptor dye. Single molecule experiments not only probe average distances, but also distance distributions or even fluctuations, and thus provide a powerful tool to study biomolecular structure and dynamics. However, the measured energy transfer efficiency depends not only on the distance between the dyes, but also on their mutual orientation, which is typically inaccessible to experiments. Read More
    Combining Graphical and Analytical Methods with Molecular Simulations To Analyze Time-Resolved FRET Measurements of Labeled Macromolecules Accurately.
    J Phys Chem B 2017 Sep 28;121(35):8211-8241. Epub 2017 Aug 28.
    Lehrstuhl für Molekulare Physikalische Chemie, Heinrich-Heine-Universität , Universitätsstraße 1, 40225 Düsseldorf, Germany.
    Förster resonance energy transfer (FRET) measurements from a donor, D, to an acceptor, A, fluorophore are frequently used in vitro and in live cells to reveal information on the structure and dynamics of DA labeled macromolecules. Accurate descriptions of FRET measurements by molecular models are complicated because the fluorophores are usually coupled to the macromolecule via flexible long linkers allowing for diffusional exchange between multiple states with different fluorescence properties caused by distinct environmental quenching, dye mobilities, and variable DA distances. It is often assumed for the analysis of fluorescence intensity decays that DA distances and D quenching are uncorrelated (homogeneous quenching by FRET) and that the exchange between distinct fluorophore states is slow (quasistatic). Read More
    The effect of Brownian motion of fluorescent probes on measuring nanoscale distances by Förster resonance energy transfer.
    J Chem Phys 2011 Jun;134(22):225102
    Department of Physics and Institute for Optical Sciences, University of Toronto, Toronto, Ontario M5S 1A7, Canada.
    Förster resonance energy transfer (FRET) is a powerful optical technique to determine intra-molecular distances. However, the dye rotational motion and the linker flexibility complicate the relationship between the measured energy transfer efficiency and the distance between the anchoring points of the dyes. In this study, we present a simple model that describes the linker and dye dynamics as diffusion on a sphere. Read More
    Accurate distance determination of nucleic acids via Förster resonance energy transfer: implications of dye linker length and rigidity.
    J Am Chem Soc 2011 Mar 3;133(8):2463-80. Epub 2011 Feb 3.
    Institut für Physikalische Chemie, Lehrstuhl für Molekulare Physikalische Chemie, Heinrich-Heine-Universität, Düsseldorf, Germany.
    In Förster resonance energy transfer (FRET) experiments, the donor (D) and acceptor (A) fluorophores are usually attached to the macromolecule of interest via long flexible linkers of up to 15 Å in length. This causes significant uncertainties in quantitative distance measurements and prevents experiments with short distances between the attachment points of the dyes due to possible dye-dye interactions. We present two approaches to overcome the above problems as demonstrated by FRET measurements for a series of dsDNA and dsRNA internally labeled with Alexa488 and Cy5 as D and A dye, respectively. Read More