Netropsin-poly(dA-dT) complex in solution: structure and dynamics of antibiotic-free base pair regions and those centered on bound netropsin.

Abstract

The biphasic duplex-to-strand transition for the netropsin.cntdot.poly(dA-dT) complex, phosphate/drug mole ratio (P/D) = 50, was investigated by high-resolution proton NMR spectroscopy at the nonexchangeable base and sugar protons in 0.1 M cacodylate solution. The NMB spectral parameters monitor the structure and dynamics of the opening of antibiotic-free base pair regions (55-65.degree.) and the opening of base regions centered on bound netropsin (90-100.degree.). The gradual addition of netropsin to poly(dA-dT) results in structural perturbations extending into the antibiotic-free pair regions that begin to level off above 0.02 antibiotic molecules per polynucleotide phosphate (P/D = 50). The NMR chemical shift parameters at the antibiotic-free base pair regions in the P/D = 50 complex suggest changes in the glycosidic torsion angles of the deoxyadenosine and thymidine residues and less pronounced changes in the base pair overlap geometries. The dissociation rates of the antibiotic-free base pair regions are at least an order of magnitude slower in the P/D = 50 netropsin.cntdot.poly(dA-dT) complex compared to related parameters for poly(dA-dT) and the P/D = 50 ethidium bromide.cntdot.poly(dA-dT) complex. There is decreased segmental mobility at the antibiotic-free strand regions in the temperature range (65-90.degree.) between the 2 transitions in the biphasic melting curve of the P/D = 50 netropsin.cntdot.poly (dA-dT) complex. Netropsin stabilizes at least 5 base pairs, with their center at its binding site.