Sequence-specific alkylation and cleavage of DNA mediated by purine motif triple helix formation.

Abstract

An N-bromoacetyl electrophile attached to the 5'-phosphate group of a purine-rich oligonucleotide affords sequence-specific alkylation of duplex DNA (at 37 degrees C, pH 7.4) through the formation of a specific purine.purine.pyrimidine triple-helical complex. In a 645 bp restriction fragment containing three consecutive guanine bases adjacent to the 3'-end of an oligonucleotide binding site, the yield of single-strand cleavage after piperidine treatment is 80% at the guanine base directly adjacent to the binding site and 88% overall. In an 837 bp restriction fragment containing two adjacent inverted repeats of the third strand binding site and a single 3'-guanine base, yields of single-strand cleavage are 87% on each strand at the 3'-guanine base. Double-strand cleavage was obtained in 61% yield at a single site in a 6.6 kbp plasmid containing the 837 bp fragment. Extension of triple helix mediated DNA alkylation from the pyrimidine to purine motif formally extends the number of sites in duplex DNA that can be cleaved in a sequence-specific and nucleotide-specific manner in good yields.