Journal
NUCLEIC ACIDS RESEARCH
Volume 37, Issue 12, Pages 3990-3995Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkp277
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Funding
- The Icelandic Research Fund [060028023]
- Eimskip Fund of the University of Iceland
- Science Institute, University of Iceland
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The cocaine aptamer is a DNA molecule that binds cocaine at the junction of three helices. The bifunctional spectroscopic probe C was incorporated independently into three different positions of the aptamer and changes in structure and dynamics upon addition of the cocaine ligand were studied. Nucleoside C contains a rigid nitroxide spin label and can be studied directly by electron paramagnetic resonance (EPR) spectroscopy and fluorescence spectroscopy after reduction of the nitroxide to yield the fluoroside C-f. Both the EPR and the fluorescence data for aptamer 2 indicate that helix III is formed before cocaine binding. Upon addition of cocaine, increased fluorescence of a fully base-paired C-f, placed at the three-way junction in helix III, was observed and is consistent with a helical tilt from a coaxial stack of helices II and III. EPR and fluorescence data clearly show that helix I is formed upon addition of cocaine, concomitant with the formation of the Y-shaped three-way helical junction. The EPR data indicate that nucleotides in helix I are more mobile than nucleotides in regular duplex regions and may reflect increased dynamics due to the short length of helix I.
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