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Gauging the flexibility of fluorescent markers for the interpretation of fluorescence resonance energy transfer

Rindermann, Jan J., Akhtman, Yosef, Richardson, James, Brown, Tom and Lagoudakis, Pavlos G. (2011) Gauging the flexibility of fluorescent markers for the interpretation of fluorescence resonance energy transfer Journal of the American Chemical Society, 133, (2), pp. 279-285. (doi:10.1021/ja105720j).

Record type: Article

Abstract

Intramolecular distances in proteins and other biomolecules can be studied in living cells by means of fluorescence resonance energy transfer (FRET) in steady-state or pulsed-excitation experiments. The major uncertainty originates from the unknown orientation between the optical dipole moments of the fluorescent markers, especially when the molecule undergoes thermal fluctuations in physiological conditions. We introduce a statistical method based on the von Mises?Fisher distribution for the interpretation of fluorescence decay dynamics in donor?acceptor FRET pairs that allows us to retrieve both the orientation and the extent of directional fluctuations of the involved dipole moments. We verify the method by applying it to donor?acceptor pairs controllably attached to DNA helices and find that common assumptions such as complete rotational freedom or fully hindered rotation of the dipoles fail a physical interpretation of the fluorescence decay dynamics. This methodology is applicable in single-molecule and ensemble measurements of FRET to derive more accurate distance estimates from optical experiments, without the need for more complex and expensive NMR studies

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Published date: 2011

Identifiers

Local EPrints ID: 179809
URI: http://eprints.soton.ac.uk/id/eprint/179809
ISSN: 0002-7863
PURE UUID: 5fdce93e-6ceb-46de-9c99-487ec7905d0f

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Date deposited: 13 Apr 2011 09:11
Last modified: 18 Jul 2017 12:02

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Contributors

Author: Jan J. Rindermann
Author: Yosef Akhtman
Author: James Richardson
Author: Tom Brown
Author: Pavlos G. Lagoudakis

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