Spectroscopic and Thermodynamic Studies on the Binding of Sanguinarine and Berberine to Triple and Double Helical DNA and RNA Structures http://www.jbsdonline.com Abstract A comparative study on the interaction of sanguinarine and berberine with DNA and RNA triplexes and their parent duplexes was performed, by using a combination of spectrophoto- metric, UV thermal melting, circular dichroic and thermodynamic techniques. Formation of the DNA and RNA triplexes was confirmed from UV-melting and circular dichroic meas- urements. The interaction process was characterized by increase of thermal melting tem- perature, perturbation in circular dichroic spectrum and the typical hypochromic and bathochromic effects in the absorption spectrum. Scatchard analysis indicated that both the alkaloids bound to the triplex and duplex structures in a non-cooperative manner and the binding was stronger to triplexes than to parent duplexes. Thermal melting studies further indicated that sanguinarine stabilized the Hoogsteen base paired third strand of both DNA and RNA triplexes more tightly compared to their Watson-Crick strands, while berberine stabilized the third strand only without affecting the Watson-Crick strand. However, san- guinarine stabilized the parent duplexes while no stabilization was observed with berberine under identical conditions. Circular dichroic studies were also consistent with the observa- tion that perturbations of DNA and RNA triplexes were more compared to their parent duplexes in presence of the alkaloids. Thermodynamic data revealed that binding of san- guinarine and berberine to triplexes (T.AxT and U.AxU) and duplexes (A.T and A.U) showed negative enthalpy changes and positive entropy changes but that of sanguinarine to C.GxC + triplex and G.C duplex exhibited negative enthalpy and negative entropy changes. Taken together, these results suggest that both sanguinarine and berberine can bind and sta- bilize the DNA and RNA triplexes more strongly than their respective parent duplexes. Introduction Triple helices were first described in 1957 (1) but interest in these structures has recently intensified due to the realization that synthetic oligonucleotide-directed triple helix formation can be used as antigene agents for selectively targeting spe- cific DNA sequences (2-4). The antigene strategy remains one of the most fasci- nating fields of triplex application to selectively control gene expression (5, 6). The formation of intermolecular DNA triplexes offers a means of achieving sequence specific recognition of long DNA sequences. In this strategy, a synthetic oligonu- cleotide binds within the major groove of duplex DNA, forming specific hydrogen bond to substituents on the DNA bases. Although the third strand oligonucleotide possesses specific sequence recognition properties, the binding is not so strong, which results in a relatively low stability of the triplex structure. The instability of triple helical structures under normal physiological conditions is a critical limitation that restricts their uses in vivo. Various strategies have thus been explored for enhancing triplex stability. This includes tethering of intercalating agents to the end of third-strand oligonucleotides (7, 8), photoinduction of covalent cross-links between the third strand and the underlying duplex via a tethered photoactivatable Journal of Biomolecular Structure & Dynamics, ISSN 0739-1102 Volume 20, Issue Number 5, (2003) ©Adenine Press (2003) Suman Das Gopinatha Suresh Kumar Arghya Ray Motilal Maiti * Biophysical Chemistry Laboratory Indian Institute of Chemical Biology 4 Raja S.C. Mullick Road Kolkata 700032, India 1 *Phone: 0091 33 472 4049 Fax : 0091 33 473 0284 / 5197 Email : mmaiti@iicb.res.in