An. Veselkov et al., H-1-NMR STUDY OF THE INTERACTION BETWEEN ETHIDIUM-BROMIDE AND SELF-COMPLEMENTARY DEOXYTETRANUCLEOTIDE 5'-D(CPGPCPG) IN AN AQUEOUS-SOLUTION, Journal of structural chemistry, 37(1), 1996, pp. 65-75
Complexation of the ethidium bromide (3,8-diamino-6-phenyl-5-ethylphen
anthridine) dye with self-complementary deoxytetraribonucleoside triph
osphate 5'-d(CpGpCpC) in a water-salt solution is studied by one- and
two-dimensional H-1 NMR spectroscopy (500 MHz). Two-dimensional PMR sp
ectroscopy (2D-COSY and 2D-NOESY) is used for the full assignment of p
roton signals of the molecules in the solution and for the qualitative
analysis of the interaction between ethidium bromide and the tetranuc
leotide. The concentration dependences of the proton chemical shifts o
f the molecules at a fired temperature (T = 308 K) are measured Differ
ent schemes of the formation of the dye complexes with the tetranucleo
tide, taking into account various molecular associations in the soluti
on are considered. The equilibrium constants of the reactions and the
limiting chemical shifts of ethidium bromide protons in the complexes
are determined. The relative contents of different complexes in the so
lution are analyzed and the dynamic equilibrium is studied as a functi
on of the dye-tetranucleotide ratio in the solution. The data obtained
suggest that ethidium bromide (like the acridine dye proflavine) is i
ntercalated predominantly into the pyrimidine-purine sections (CG site
s) of the tetranucleotide duplex. However, ethidium bromide is interca
lated an the side of the narrow slot of the duplex, while proflavine i
ntercalation occurs through the broad slot of the double helix. The mo
st likely structures of the 1:2 and 2:2 dye-tetranucleotide complexes
in the solution are constructed using the calculated values of the ind
uced chemical shifts of ethidium bromide protons and 2D NOESY spectros
copy data.