Biopolym. Cell. 2004; 20(3):215-223.
Structure and Function of Biopolymers
Thermodynamics of complexation of aromatic ligand with single-stranded deoxyoligonucleotide and a hairpin loop of the same nucleotide content
1Baranovsky S. F., 1Rogova O. V., 2Hernandez Santiago A. A., 1Zavyalova O. S., 1Veselkov A. N.
  1. Sevastopol National Technical University
    33, Universytetska Str., Sevastopol, Ukraine, 99053
  2. Autonomous University of Puebla
    Puebla, Mexico, 72570


Comparative analysis of the thermodynamical parameters of interaction of phenanthridine dye, ethidium bromide (EB) with deoxyheptanucleotide d(GpCpGpApApGpC), capable to form a hairpin structure, and a single-stranded deoxytetranucleotide d(GpApApG), being a component element of the heptamer sequence, has been made using 1NMR spectroscopy (500/600 MHz). Thermodynamical parameters (ΔH, DS) of complexation of EB with oligonucleotides have been calculated from the experimental temperature dependences of proton chemical shifts using a model of dynamic equilibrium between different conformational states of oligomers in solution. For example, monomer, hairpin and dimer forms have been considered as the heptamer conformational states in aqueous solution. The calculated enthalpy of complexation of EB with single-stranded tetranucleotide d(GAAG) is similar, within the error limits, to the value of complex formation between the dye molecule and the hairpin loop d(GAA). Analysis of the results obtained enables to conclude that single-stranded non-self-complementery oligonucleotides of the same nucleotide content as in the hairpin loop may be used as model systems for the investigation of complexation of aromatic ligands with hairpin structures in solution.


[1] Bianchi ME, Beltrame M, Paonessa G. Specific recognition of cruciform DNA by nuclear protein HMG1. Science. 1989;243(4894 Pt 1):1056-9.
[2] Duckett DR, Murchie AI, Bhattacharyya A, Clegg RM, Diekmann S, von Kitzing E, Lilley DM. The structure of DNA junctions and their interaction with enzymes. Eur J Biochem. 1993;211(1-2):285-95.
[3] Spiro C, Richards JP, Chandrasekaran S, Brennan RG, McMurray CT. Secondary structure creates mismatched base pairs required for high-affinity binding of cAMP response element-binding protein to the human enkephalin enhancer. Proc Natl Acad Sci U S A. 1993;90(10):4606-10.
[4] Miao DM, Honda Y, Tanaka K, Higashi A, Nakamura T, Taguchi Y, Sakai H, Komano T, Bagdasarian M. A base-paired hairpin structure essential for the functional priming signal for DNA replication of the broad host range plasmid RSF1010. Nucleic Acids Res. 1993;21(21):4900-3.
[5] Froelich-Ammon SJ, Gale KC, Osheroff N. Site-specific cleavage of a DNA hairpin by topoisomerase II. DNA secondary structure as a determinant of enzyme recognition/cleavage. J Biol Chem. 1994;269(10):7719-25.
[6] Chou SH, Zhu L, Gao Z, Cheng JW, Reid BR. Hairpin loops consisting of single adenine residues closed by sheared A.A and G.G pairs formed by the DNA triplets AAA and GAG: solution structure of the d(GTACAAAGTAC) hairpin. J Mol Biol. 1996;264(5):981-1001.
[7] Brown DR, Kurz M, Kearns DR, Hsu VL. Formation of multiple complexes between actinomycin D and a DNA haiBiochemistry. 1994;33(3):651-64.rpin: structural characterization by multinuclear NMR.
[8] Davies DB, Pahomov VI, Veselkov AN. NMR determination of the conformational and drug binding properties of the DNA heptamer d(GpCpGpApApGpC) in aqueous solution. Nucleic Acids Res. 1997;25(22):4523-31.
[9] Veselkov AN, Pakhomov VI, Baranovskii SF, Davies DB. Analysis of conformation of oligonucleotide with sequences 5'-d(GpCpGpApApGpC) by 1H-NMR-spectroscopy. Mol Biol (Mosk). 1997;31(6):1036-42.
[10] Rentzeperis D, Medero M, Marky LA. Thermodynamic investigation of the association of ethidium, propidium and bis-ethidium to DNA hairpins. Bioorg Med Chem. 1995;3(6):751-9.
[11] Hirao I, Kawai G, Yoshizawa S, Nishimura Y, Ishido Y, Watanabe K, Miura K. Most compact hairpin-turn structure exerted by a short DNA fragment, d(GCGAAGC) in solution: an extraordinarily stable structure resistant to nucleases and heat. Nucleic Acids Res. 1994;22(4):576-82.
[12] Reinhardt CG, Krugh TR. A comparative study of ethidium bromide complexes with dinucleotides and DNA: direct evidence for intercalation and nucleic acid sequence preferences. Biochemistry. 1978;17(23):4845-54.
[13] Altona C, Van Boom JH, Haasnoot CA. Conformational analysis of a DNA triplet in aqueous solution. Thymidylyl-(3'-5')-thymidylyl-(3'-5')-2'-deoxyadenosine, d(T-T-A), studied by 1H nuclear magnetic resonance at 360 MHz. Eur J Biochem. 1976;71(2):557-62.
[14] Veselkov A. N., Baranovsky S. F., Petrenko N. V., Osetrov S. G., Veselkov D. A., Djimant L. N., Tucker A., Parkes H., Davies D. 1H-NMR investigation of the self-association of non-complementary deoxytetranucleotides of different base sequences in aqueous solution. Biopolym Cell. 1996; 12(4):38-48.
[15] Veselkov AN, Dymant LN, Baranovskiy SF, Bolotin PA, Parkes XYe, Devis D. Investigation of self-association of ethidium bromide in aqueous solution by 1H-NMR spectroscopy. Khim fizika. 1994; 13:70-8.
[16] Davies D.B., Veselkov A.N. Structural and thermodynamical analysis of molecular complexation by 1H NMR spectroscopy: Intercalation of ethidium bromide with the isomeric deoxytetranucleoside triphosphates 5'-d(GpCpGpC) and 5'-d(CpGpCpG) in aqueous solution. Journal of the Chemical Society - Faraday Transactions, 1996; 92 (19):3545-3557.
[17] Davies D.B., Baranovsky S.F., Veselko, A.N. Structural and thermodynamical analysis of drug binding to single-stranded DNA oligomers: Self-association of non-self-complementary deoxytetranucleotides of different base sequence and their complexation with ethidium bromide in aqueous solution. Journal of the Chemical Society - Faraday Transactions, 1997; 93 (8):1559-1572.
[18] Veselkov A.N., Djimant L.N., Karawajew L.S., Kulikov E.L. Investigation of the aggregation of acridine dyes in aqueous solution by 'H NMR. Stud biophys. 1985;120:171-180.
[19] Davies DB, Djimant LN, Baranovsky SF, Veselkov AN. 1H-NMR determination of the thermodynamics of drug complexation with single-stranded and double-stranded oligonucleotides in solution: ethidium bromide complexation with the deoxytetranucleotides 5'-d(ApCpGpT), 5'-d(ApGpCpT), and 5'-d(TpGpCpA). Biopolymers. 1997;42(3):285-95.
[20] Rentzeperis D, Alessi K, Marky LA. Thermodynamics of DNA hairpins: contribution of loop size to hairpin stability and ethidium binding. Nucleic Acids Res. 1993;21(11):2683-9.
[21] Chaires JB. Thermodynamics of the daunomycin-DNA interaction: ionic strength dependence of the enthalpy and entropy. Biopolymers. 1985;24(2):403-19.