Bleomycin-A2 complexes with poly(dA-dT): A proton nuclear magnetic resonance study of the nonexchangeable hydrogens

Binding of bleomycin-A2 (Bleo-A2) to poly(dA-dT) in 100 mM sodium phosphate (pH 6.8) in D2O has been investigated by 1H NMR spectroscopy at 270 and 360 MHz. Significant spectral perturbations were observed only when the nucleic acid was in the duplex state. Of the Bleo-A2 resonances, the two bithiazole peaks exhibited the largest spectral shiffs and line broadening effects. The high field shift of these resonances was very small near room temperature and reached a maximum of about 0.27 ppm just below the thermal denaturation temperature of the nucleic acid (Tm = 60 ± 1°C). The temperature dependence of spectral perturbations may be accounted for by the formation of at least two types of Bleo-A2 complexes with the polynucleotide. Other perturbed resonances of bleomycin are the S-C H 3 and S-C H 2 of the terminal amine, the C H 2-N resonance of the bithiazole residue, and the C H (CH3)CO of the methylvaleric acid residue. The significantly perturbed resonances of the nucleic acid originate from the A(H-2), A(H-8), T(H-6) and one of the H-2′ hydrogens. Binding of the C-terminal tripeptide fragment of Bleo-A2 to poly(dA-dT) is accompanied by selective broadening of the bithiazole group. These experiments have identified potential loci of interaction on the Bleo-A2 and poly(dA-dT) molecules.