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Cyanine Dyes as Intercalating Agents: Kinetic and Thermodynamic Studies on the DNA/Cyan40 and DNA/CCyan2 Systems

Tarita Biver ^*, Angela De Biasi ^*, Fernando Secco ^*, Marcella Venturini ^* and Sergiy Yarmoluk 

^* Dipartimento di Chimica e Chimica Industriale, Università di Pisa, Pisa, Italy; and Institute of Molecular Biology and Genetics of the National Academy of Sciences of Ukraine, Kyiv, Ukraine

Correspondence: Address reprint requests to Fernando Secco, Dipartimento di Chimica e Chimica Industriale, Via Risorgimento, 35-56126 Pisa, Italy. Tel.: 0039-050-221-9259; Fax: 0039-050-221-9260; E-mail: ferdi{at}dcci.unipi.it.

The interaction of cyanines with nucleic acids is accompanied by intense changes of their optical properties. Consequently these molecules find numerous applications in biology and medicine. Since no detailed information on the binding mechanism of DNA/cyanine systems is available, a T-jump investigation of the kinetics and equilibria of binding of the cyanines Cyan40 [3-methyl-2-(1,2,6-trimethyl-4(1H)pyridinylidenmethyl)-benzothiazolium ion] and CCyan2 [3-methyl-2-[2-methyl-3-(3-methyl-2(3H)-benzothiazolylidene)-1-propenyl]-benzothiazolium ion] with CT-DNA is performed at 25°C, pH 7 and various ionic strengths. Bathochromic shifts of the dye absorption band upon DNA addition, polymer melting point displacement (T = 8–10°C), site size determination (n = 2), and stepwise kinetics concur in suggesting that the investigated cyanines bind to CT-DNA primary by intercalation. Measurements with poly(dA-dT)·poly(dA-dT) and poly(dG-dC)·poly(dG-dC) reveal fair selectivity of CCyan2 toward G-C basepairs. T-jump experiments show two kinetic effects for both systems. The binding process is discussed in terms of the sequence D + S D,S DS_I DS_II, which leads first to fast formation of an external complex D,S and then to a partially intercalated complex DS_I which, in turn, converts to DS_II, a more stable intercalate. Absorption spectra reveal that both dyes tend to self-aggregate; the kinetics of CCyan2 self-aggregation is studied by T-jump relaxation and the results are interpreted in terms of dimer formation.