Drug Binding by Branched DNA Molecules: Analysis by Chemical Footprinting of Intercalation into an Immobile Junction

Branched DNA structures interact with drugs differently from unbranched control duplexes of similar sequence. A specific interaction between the reagent (methidiumpropyl-EDTA)-Fe(II) [MPE-Fe(II)] and a branched DNA molecule formed from 16-mer oligonucleotide strands has been reported [Guo, Q., Seeman, N. C, & Kallenbach, N. R. (1989) Biochemistry 28, 2355–2359]. The structure of the branched molecule is thought to be made up of two double-helical stacking domains with an overall twofold symmetry across the branch site. The MPE-Fe(II) interaction occurs predominantly at or adjacent to the branch site and is eliminated by a second intercalator, propidium iodide. Further studies on the nature and properties of this site are presented here. Comparison of the patterns of scission of linear duplex and branched tetramer by EDTA-Fe(II), MPE-Fe(II), and Cu(I)-(o-phenanthroline)₂ [(OP)₂Cu(I)] provides a higher resolution picture of the site of enhanced binding. In particular, the sensitive footprinting afforded by (OP)₂Cu(I) allows us to localize the major site of preferential interaction with propidium precisely to the branch point itself, with a roughly twofold symmetric pattern of cuts resulting. In detail, the differential pattern with respect to each duplex control is distinct for each arm of the junction. Excess propidium results in apparent reversal of the crossover isomer of the junction, indicating a possible additional avenue for the action of drugs in biological systems-effects on the products of recombination.