Estimation of the diversity between DNA calorimetric profiles, differential melting curves and corresponding melting temperatures.

The Poland-Fixman-Freire formalism was adapted for modeling of calorimetric DNA melting profiles, and applied to plasmid pBR 322 and long random sequences. We studied the influence of the difference (HGC -HAT ) between the helix-coil transition enthalpies of AT and GC base pairs on the calorimetric melting profile and on normalized calorimetric melting profile. A strong alteration of DNA calorimetrical profile with HGC -HAT was demonstrated. In contrast, there is a relatively slight change in the normalized profiles and in corresponding ordinary (optical) normalized differential melting curves (DMCs). For fixed HGC -HAT , the average relative deviation (S) between DMC and normalized calorimetric profile, and the difference between their melting temperatures (Tcal -Tm ) are weakly dependent on peculiarities of the multipeak fine structure of DMCs. At the same time, both the deviation S and difference (Tcal -Tm ) enlarge with the temperature melting range of the helix-coil transition. It is shown that the local deviation between DMC and normalized calorimetric profile increases in regions of narrow peaks distant from the melting temperature.

Determination of melting temperature and temperature melting range for DNA with multi-peak differential melting curves.

Many factors that change the temperature position and interval of the DNA helix-coil transition often also alter the shape of multi-peak differential melting curves (DMCs). For DNAs with a multi-peak DMC, there is no agreement on the most useful definition for the melting temperature, Tm, and temperature melting width, ΔT, of the entire DNA transition. Changes in Tm and ΔT can reflect unstable variation of the shape of the DMC as well as alterations in DNA thermal stability and heterogeneity. Here, experiments and computer modeling for DNA multi-peak DMCs varying under different factors allowed testing of several methods of defining Tm and ΔT. Indeed, some of the methods give unreasonable "jagged" Tm and ΔT dependences on varying relative concentration of DNA chemical modifications (rb), [Na(+)], and GC content. At the same time, Tm determined as the helix-coil transition average temperature, and ΔT, which is proportional to the average absolute temperature deviation from this temperature, are suitable to characterize multi-peak DMCs. They give smoothly varying theoretical and experimental dependences of Tm and ΔT on rb, [Na(+)], and GC content. For multi-peak DMCs, Tm value determined in this way is the closest to the thermodynamic melting temperature (the helix-coil transition enthalpy/entropy ratio).

Comparative thermal and thermodynamic study of DNA chemically modified with antitumor drug cisplatin and its inactive analog transplatin.

Antitumor activity of cisplatin is exerted by covalent binding to DNA. For comparison, studies of cisplatin-DNA complexes often employ the very similar but inactive transplatin. In this work, thermal and thermodynamic properties of DNA complexes with these compounds were studied using differential scanning calorimetry (DSC) and computer modeling. DSC demonstrates that cisplatin decreases thermal stability (melting temperature, Tm) of long DNA, and transplatin increases it. At the same time, both compounds decrease the enthalpy and entropy of the helix-coil transition, and the impact of transplatin is much higher. From Pt/nucleotide molar ratio rb=0.001, both compounds destroy the fine structure of DSC profile and increase the temperature melting range (ΔT). For cisplatin and transplatin, the dependences δTm vs rb differ in sign, while δΔT vs rb are positive for both compounds. The change in the parameter δΔT vs rb demonstrates the GC specificity in the location of DNA distortions. Our experimental results and calculations show that 1) in contrast to [Pt(dien)Cl]Cl, monofunctional adducts formed by transplatin decrease the thermal stability of long DNA at [Na(+)]>30mM; 2) interstrand crosslinks of cisplatin and transplatin only slightly increase Tm; 3) the difference in thermal stability of DNA complexes with cisplatin vs DNA complexes with transplatin mainly arises from the different thermodynamic properties of their intrastrand crosslinks. This type of crosslink appears to be responsible for the antitumor activity of cisplatin. At any [Na(+)] from interval 10-210mM, cisplatin and transplatin intrastrand crosslinks give rise to destabilization and stabilization, respectively.