Footprinting with MPE.Fe(II). Complementary-strand analyses of distamycin- and actinomycin-binding sites on heterogeneous DNA.

Abstract

We recently reported a direct technique for determining the binding sites of small molecules on naturally occurring heterogeneous DNA (Van Dyke et al. 1982). Methidiumpropyl-EDTA·Fe(II) (MPE·Fe[II]) (Hertzberg and Dervan 1982) cleaves double-helical DNA with low sequence-specificity (Van Dyke et al. 1982). Using a combination of MPE·Fe(II) partial cleavage of drug-protected DNA fragments and Maxam-Gilbert sequencing methods, we determined the drug-protected sites on one strand of a double-helical fragment from pBR322 for the intercalator actinomycin D (Goldberg et al. 1962; Muller and Crothers 1968; Wells and Larson 1970; Sobell 1973; Krugh 1981; Patel et al. 1981; Takusagawa et al. 1982) and the minor-groove binders netropsin and distamycin A (Luck et al. 1974; Wartell et al. 1974; Zimmer 1975; Berman et al. 1979; Krylov et al. 1979). Netropsin and distamycin A gave identical DNA-cleavage inhibition patterns or footprints in regions rich in dA·dT base pairs. Actinomycin D afforded a completely different footprint...