THE BINDING OF AN ANTISENSE OLIGONUCLEOTIDE TO A HAIRPIN STRUCTURE VIA TRIPLEX FORMATION INHIBITS CHEMICAL AND BIOLOGICAL REACTIONS

We have investigated the binding of a 26-mer antisense oligodeoxynucle otide to a 69-mer DNA hairpin with a 13 base pair stem, bearing an Rsa 1 restriction site. The 5' part of the 26-mer annealed to a stretch of six purines at the bottom of the hairpin. The 3' part was designed to fold back to form a triplex with both the stem of the hairpin and wit h the sequence paired to its own 5' region. Using non-denaturing polya crylamide gel electrophoresis, melting curves (Tm) and chemical footpr inting, we were able to show the formation of a 'double-hairpin' compl ex between the 69-mer and the 26-mer antisense oligopyrimidines. The a ssociation was both sequence and pH-dependent. The formation of a doub le hairpin complex was shown to prevent the alkylation of the 69-mer D NA target by an oligonucleotide-nitrogen mustard conjugate and to sele ctively inhibit the action of Rsa1.