Reevaluation of a dna binding activity with specificity for chemically modified dna

A DNA binding activity which appeared in direct filter binding assays to show specificity for DNA modified by N-acetoxy-2-acetylaminofluorene (AAF), N-methyl-N-nitro-sourea and methylmethanesulfonate (Moranelli and Lieber-man, Proc. Natl. Acad. Sci. USA, 77, 3201, 1980) has been further purified and characterized. Equilibrium competition binding experiments indicate this activity shows little, or no, preferential binding of AAF-modified DNA compared to single-stranded DNA and only a small preference compared to u.v.-irradiated DNA or double-stranded DNA. These findings demonstrate the difficulty of using direct filter binding assays when purifying proteins with a desired biological function and the need to perform competition assays when evaluating specificity.