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Salt Dependence, Thermodynamic Parameters and Thermal Denaturation of the ct-DNA Binding of Methyl Green
Submitted by redaksi on Fri, 09/24/2010 - 05:36.
* Corresponding authors: Phone/Fax: +62 274 545188; E-mail: mudasir(at)ugm.ac.id or dasir01(at)yahoo.com
Mudasir*,1, Endang Tri Wahyuni1, Nurul H. Aprillita1, Naoki Yoshioka2 and Hidenari Inoue2
1Department of Chemistry, Faculty of Mathematics and Natural Sciences, Gadjah Mada University, Sekip Utara, P.O. Box Bls. 21, Yogyakarta 55281, Indonesia
2Department of Applied Chemistry, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama 223-8522, Japan
* Corresponding authors: Phone/Fax: +62 274 545188; E-mail: mudasir(at)ugm.ac.id or dasir01(at)yahoo.com
Abstract
The ct-DNA binding properties of methyl green (MG) including salt dependence of the binding constant, thermodynamic parameters and thermal denaturation change (Tm) have been systematically studied by spectrophotometric titration method. The binding of MG to ct-DNA is quite strong as indicated by hypochromic effect at high ratio of [dye]/[ct-DNA] (R) followed by hyperchromic effect at lower ratio of R, red shift of 10 nm and the equilibrium binding constant (Kb) in the order of 105 M-1 bp. Salt-dependent experiment revealed that the Kb value of the ct-DNA binding of MG decreases with the increase of salt concentration in the medium, suggesting that in addition to intercalation process, electrostatic interaction also operate in the binding event, especially at lower ratio of R. However, the charge release (Z) by ct-DNA upon interaction with MG is somewhat lower, e.g. +0.632 at high R and +0.835 at low R as compared to positive charge carried by the MG dye. Van Hoff plot of 1/T versus ln Kb at various temperatures suggests that the cationic MG dye binds endothermically to ct-DNA, indicating that the ct-DNA binding of MG is entropically driven. Thermal denaturation experiments suggest that a remarkable stabilization of double helix ct-DNA is observed when the MG dye binds to ct-DNA as indicated by the increase in Tm of ct-DNA of about 12 oC in the presence of MG. This value of Tm change upon binding of MG to ct-DNA is comparable to those observed for common intercalators such as ethidium bromide and daunomycin, suggesting that MG most likely binds to ct-DNA via both electrostatic and intercalation binding modes.
Keywords: Methyl green, DNA, thermodynamic parameter, cationic dye, binding constant