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Journal of Chemical Physics / Volume 134 / Issue 22 / ARTICLES / Biological Molecules, Biopolymers, and Biological Systems

J. Chem. Phys. 134, 225102 (2011); http://dx.doi.org/10.1063/1.3598109 (11 pages)

The effect of Brownian motion of fluorescent probes on measuring nanoscale distances by Förster resonance energy transfer

Daniel Badali and Claudiu C. Gradinaru

Department of Physics and Institute for Optical Sciences, University of Toronto, Toronto, Ontario M5S 1A7, Canada and Department of Chemical and Physical Sciences, University of Toronto Mississauga, Mississauga, Ontario L5L 1C6, Canada

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(Received 16 December 2010; accepted 17 May 2011; published online 9 June 2011)

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. Single-pair energy transfer was treated in the weak excitation limit, photon statistics and scaffold flexibility were ignored, and different time-averaging regimes were considered. Despite the approximations, our model provides new insights for experimental designs and results interpretation in single-molecule FRET. Monte Carlo simulations produced distributions of the inter-dye distance, the dipole orientation factor, κ2, and the transfer efficiency, E, which were in perfect agreement with independently derived theoretical functions. Contrary to common perceptions, our data show that longer linkers will actually restrict the motion of dye dipoles and hence worsen the isotropic 2/3 approximation of κ2. It is also found that the thermal motions of the dye-linker system cause fast and large efficiency fluctuations, as shown by the simulated FRET time-trajectories binned on a microsecond time scale. A fundamental resolution limit of single-molecule FRET measurements emerges around 1–10 μs, which should be considered for the interpretation of data recorded on such fast time scales.

© 2011 American Institute of Physics

Article Outline

  1. INTRODUCTION
  2. METHODS
  3. RESULTS AND DISCUSSION
    1. Inter-dye distance distribution
    2. κ 2 distributions
    3. FRET efficiency distributions
    4. Brownian noise
  4. CONCLUSIONS

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KEYWORDS and PACS

Keywords

Brownian motion, diffusion, dyes, excited states, fluorescence, intramolecular forces, Monte Carlo methods, rotational states

PACS

  • 34.20.Gj

    Intermolecular and atom-molecule potentials and forces

  • 02.70.Uu

    Applications of Monte Carlo methods

  • 05.40.Jc

    Brownian motion

  • 33.50.Dq

    Fluorescence and phosphorescence spectra

ARTICLE DATA

Digital Object Identifier
http://dx.doi.org/10.1063/1.3598109

PUBLICATION DATA

ISSN

0021-9606 (print)  
1089-7690 (online)

Publisher

$abstract.society.value is a Member of CrossRef American Institute of Physics

For access to fully linked references, you need to log in.
    B. A. Camley, F. L. H. Brown, and E. A. Lipman, J. Chem. Phys. 131(10) (2009)JCPSA6000131000010104509000001.

    I. Gopich and A. Szabo, J. Chem. Phys. 122(1) (2005)JCPSA6000122000001014707000001.

    D. E. Makarov and K. W. Plaxco, J. Chem. Phys. 131(8), 085105 (2009)JCPSA6000131000008085105000001.


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