[1H-NMR study of complex formation between the aromatic dye ethidium bromide and the single-stranded noncomplementary deoxytetranucleotide 5'-d(CpTpGpA) in aqueous solution].
Complex formation between the aromatic dye ethidium bromide (3,8-diamino-6-phenyl-5-ethylphenanthridine) and the single-stranded noncomplementary deoxytetranucleotide 5'-d(CpTpGpA) in aqueous solution was studied by one- and two-dimensional 1H NMR spectroscopy (500 and 600 MHz). Complete assignments of proton signals from the deoxytetranucleotide 5'-d(CTGA) were made using 2D-TOCSY and 2D-NOESY spectra. The concentration dependences of proton chemical shifts of the tetranucleotide were measured at T = 298 K to determine the self-association constants of tetranucleotide molecules in solution. Due to a small probability of tetranucleotide duplex formation, the complexing of dye molecules with the monomer of the tetramer plays the main role in the equilibrium of the complex in solution, which makes it possible to analyze the specificity of interactions of the aromatic ligand with the single-stranded DNA. Different schemes of complex formation were examined, and equilibrium constants for the reactions and the limiting chemical shifts of dye protons in different complexes were determined. The relative content of different types of complexes was analyzed, and specific features of the dynamic equilibrium were revealed as a function of the dye-tetranucleotide concentration ratio. The analysis indicates a sequence-specific binding of ethidium bromide to the single-stranded oligonucleotide. The structures of 1:1 complexes of the dye with the single strand of the tetranucleotide that correspond to two most probable orientations of the dye chromophore in the pyrimidine-purine T-G-site of the tetramer base sequence were constructed using the calculated values of induced chemical shifts of ethidium bromide protons.
Veselkov
A N
AN
Sevastopol Technical University, Studgorodok, Ukraine.
Baranovskiĭ
S F
SF
Dymant
L N
LN
Petrenko
N V
NV
Osetrov
S G
SG
Veselkov
D A
DA
Parkes
X
X
Davies
D
D
D001665:Binding Sites; D004996:Ethidium / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D009838:Oligodeoxyribonucleotides / Q000737:chemistry*
D005456:Fluorescent Dyes; D009838:Oligodeoxyribonucleotides; D004996:Ethidium
Issledovaniie kompleksoobrazovaniia aromaticheskogo krasitelia bromistogo etidiia s odnotsepochechnym nekomplementarnym dezoksi tetranucleotidom 5'-d(CpTpGpA) v vodnom rastvore metodom 1H IaMR -spektroskopii.