G-Quadruplex DNA pp 147-158

Part of the Methods in Molecular Biology book series (MIMB, volume 608)

Monitoring the Temperature Unfolding of G-Quadruplexes by UV and Circular Dichroism Spectroscopies and Calorimetry Techniques

Protocol

Abstract

DNA oligonucleotides containing guanine repeat sequences can adopt G-quadruplex (GQ) structures in the presence of specific metal ions. We report on how to use a combination of spectroscopic and calorimetric techniques to determine the spectral characteristics and thermodynamic parameters for the temperature-unfolding of GQs. Specifically, we investigated the unfolding of d(G2T2G2TGTG2T2G2), G2, and d(G3T2G3TGTG3T2G3), G3 by a combination of UV and circular dichroism (CD) spectroscopies, and differential scanning calorimetry (DSC).

Analysis of the UV and CD spectra of these GQs at low (100% helix) and high (100% random coil) temperatures yielded the optimal wavelengths to determine the melting curves. In addition, the CD spectra yielded the particular conformation(s) that each GQ adopted at low temperature. DSC curves yielded complete thermodynamic profiles for the unfolding of each GQ. We use these profiles to determine the thermodynamic contributions for the formation of a G-quartet stack.

Key words

Thrombin-binding aptamer G-quartets DNA quadruplexes Cations UV and CD melting curves DSC thermograms Thermodynamics Heat and stability 

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2010

Authors and Affiliations

  1. 1.Department of Pharmaceutical SciencesUniversity of Nebraska Medical CenterOmahaUSA
  2. 2.Department of Pharmaceutical Sciences, Department of Biochemistry and Molecular BiologyEppley Institute for Research in Cancer, University of Nebraska Medical CenterOmahaUSA

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