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Orientational Dynamics and Dye-DNA Interactions in a Dye-Labeled DNA Aptamer
Wilson, George S. Unruh, Jay R. Gokulrangan, Giridharan Lushington, Gerald H. Johnson, Carey K.
Wilson, George S.
Unruh, Jay R.
Gokulrangan, Giridharan
Lushington, Gerald H.
Johnson, Carey K.
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Abstract
We report the picosecond and nanosecond timescale rotational dynamics of a dye-labeled DNA oligonucleotide
or ‘‘aptamer’’ designed to bind specifically to immunoglobulin E. Rotational dynamics in combination with fluorescence lifetime
measurements provide information about dye-DNA interactions. Comparison of Texas Red (TR), fluorescein, and tetramethylrhodamine
(TAMRA)-labeled aptamers reveals surprising differences with significant implications for biophysical studies
employing such conjugates. Time-resolved anisotropy studies demonstrate that the TR- and TAMRA-aptamer anisotropy
decays are dominated by the overall rotation of the aptamer, whereas the fluorescein-aptamer anisotropy decay displays
a subnanosecond rotational correlation time much shorter than that expected for the overall rotation of the aptamer. Docking
and molecular dynamics simulations suggest that the low mobility of TR is a result of binding in the groove of the DNA helix.
Additionally, associated anisotropy analysis of the TAMRA-aptamer reveals both quenched and unquenched states that experience
significant coupling to the DNA motion. Therefore, quenching of TAMRA by guanosine must depend on the configuration
of the dye bound to the DNA. The strong coupling of TR to the rotational dynamics of the DNA aptamer, together with the
absence of quenching of its fluorescence by DNA, makes it a good probe of DNA orientational dynamics. The understanding of
the nature of dye-DNA interactions provides the basis for the development of bioconjugates optimized for specific biophysical
measurements and is important for the sensitivity of anisotropy-based DNA-protein interaction studies employing such
conjugates.
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2005-05
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Biophysical Society
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J.R. Unruh, G. Gokulrangan, G.H. Lushington, C.K. Johnson, and G.S. Wilson, Orientational Dynamics and Dye-DNA Interactions in a DNA Aptamer, Biophysical Journal. 88, 3455-3465 (2005). http://dx.doi.org/10.1529/biophysj.104.054148