Divalent metal ion effect on helix–coil transition of high molecular weight DNA in neutral and alkaline solutions

DNA conformational equilibrium in the presence of Zn2+ ions in neutral and alkaline solutions

“…The pH of the DNA solutions was 8.5 and did not change by more than 0.03 pH units upon zinc chloride addition up to 6 × 10 −4 M Zn 2+ . The DNA phosphorous concentration (P) was (2.0 ± 0.1) × 10 −5 M as determined by the molar extinction coefficient (ε m = 6600 M −1 cm −1 ) at m = 38,500 cm −1 ( m = 260 nm) [11]. The pH of the solutions was determined by a pH-meter pH-340 (Russia) with an error of ± 0.03 pH units.…”

“…Differential UV (DUV) spectra A( ) were measured at room temperature T o = 25 ± 2 • C with a UV-vis spectrophotometer (Carl Zeiss, Jena, Germany) using the four-cuvette scheme [11,13]. The spectra were normalized to DNA concentration as follows:…”

“…DNA melting curves were measured at = m = 38,500 cm −1 ( m = 260 nm) with the same spectrophotometer as was used for recording DUV spectra but employing the double-cuvette scheme [11]. After 180 min of the initial incubation at T = T o the reference cuvette was thermostated at T = T o ± 0.5 • C, while the sample cuvette was slowly heated at a rate of 0.25 • C/min to ensure the thermodynamic equilibrium of the heated solution.…”

“…In DNA B-form, N7 atoms of purine bases are the most sterically accessible from all the nitrogen base atoms; they also have the maximum molecular electrostatic potential [ [11] and references therein, [14,15]]. Interaction of different cations, including d-ions of Mt 2+ (these are the ions with d-electrons in outer orbitals) with N7 atoms of purines has been reliably documented by various techniques [14,16].…”

“…To achieve this, a dependence of the conformational transition temperature on the zinc concentration must be obtained for the whole temperature range, in which the DNA double helix is stable. Another task is to experimentally decompose the zinc-induced changes of the DNA helix-coil transition temperature (T m ) into components according to the types of DNA-metal ion interactions [11], namely coordination with N7 sites of purines or formation of metallic intramolecular bridges in AT and GC pairs [1]. The latter can be done by obtaining the concentration relationships of T m at different experimental conditions.…”