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Nucleic Acids Research 2005 33(13):4065-4077; doi:10.1093/nar/gki716
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Published online 21 July 2005

© The Author 2005. Published by Oxford University Press. All rights reserved
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Article

Length-dependent energetics of (CTG)n and (CAG)n trinucleotide repeats

Samir Amrane, Barbara Saccà, Martin Mills1, Madhu Chauhan1, Horst H. Klump1 and Jean-Louis Mergny*

Laboratoire de Biophysique, Muséum National d'Histoire Naturelle USM 503 INSERM UR 565, CNRS UMR 8646, 43 rue Cuvier, 75231 Paris cedex 05, France 1Department of Molecular and Cell Biology, University of Cape Town P.B. Rondebosh 7701, Republic of South Africa

*To whom correspondence should be addressed. Tel: +33 1 40 79 36 89; Fax: +33 1 40 79 37 05; Email: faucon{at}mnhn.fr

Received February 28, 2005. Revised June 30, 2005. Accepted June 30, 2005.

Trinucleotide repeats are involved in a number of debilitating diseases such as myotonic dystrophy. Twelve to seventy-five base-long (CTG)n oligodeoxynucleotides were analysed using a combination of biophysical [UV-absorbance, circular dichroism and differential scanning calorimetry (DSC)] and biochemical methods (non-denaturing gel electrophoresis and enzymatic footprinting). All oligomers formed stable intramolecular structures under near physiological conditions with a melting temperature that was only weakly dependent on oligomer length. Thermodynamic analysis of the denaturation process by UV-melting and calorimetric experiments revealed an unprecedented length-dependent discrepancy between the enthalpy values deduced from model-dependent (UV-melting) and model-independent (calorimetry) experiments. Evidence for non-zero molar heat capacity changes was also derived from the analysis of the Arrhenius plots and DSC profiles. Such behaviour is analysed in the framework of an intramolecular ‘branched-hairpin’ model, in which long CTG oligomers do not fold into a simple long hairpin–stem intramolecular structure, but allow the formation of several independent folding units of unequal stability. We demonstrate that, for sequences ranging from 12 to 25 CTG repeats, an intramolecular structure with two loops is formed which we will call ‘bis-hairpin’. Similar results were also found for CAG oligomers, suggesting that this observation may be extended to various trinucleotide repeats-containing sequences.

Present address: Barbara Saccà, Laboratoire de Stabilité des Génomes, Institut Pasteur, 25 rue du Dr Roux, 75724 Paris cedex 15, France


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