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Nucleic Acids Research, 1993, Vol. 21, No. 24 5616-5622
© 1993

MOLECULAR BIOLOGY

The binding of an antisense oligonucleotide to a hairpin structure via triplex formation inhibits chemical and biological reactions

Elena Brossalina+, Emanuelle Pascolo and Jean-Jacqueés Toulme*

Laboratoire de Biophysique Moleéculaire, INSERM CJF 90-13, Universiteé de Bordeaux I 146 rue Leéo Saignat, 33076 Bordeaux Cedex, France

*To whom correspondence should be addressed

Received September 13, 1993. Revised November 8, 1993. Accepted November 8, 1993.

We have investigated the binding of a 26-mer antisense oligodeoxynucleotide to a 69-mer DNA hairpin with a 13 base pair stem, bearing an Rsa1 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 with the sequence paired to its own 5' region. Using non-denaturing polyacrylamide gel electrophoresis, melting curves (Tm) and chemical footprinting, we were able to show the formation of a 'double-hairpin' complex between the 69-mer and the 26-mer antisense oligopyrimidines. The association was both sequence and pH-dependent. The formation of a double hairpin complex was shown to prevent the alkylation of the 69-mer DNA target by an oligonucleotide-nitrogen mustard conjugate and to selectively inhibit the action of Rsa1.

+Present address: Institute of Immunology, Novosibirsk, Russia


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