CHARACTERISTICS OF TRIPLER-DIRECTED PHOTOADDUCT FORMATION BY PSORALEN-LINKED OLIGODEOXYNUCLEOTIDES

A tripler-forming oligopyrimidine has been attached at its 5'-end to a photoreactive psoralen derivative and used to target a sequence which forms part of the coding region of the human aromatase gene. The 20 b ase pair sequence is not a perfect tripler target since it contains th ree pyrimidine interruptions within the purine-rich strand, Despite th is, we have detected tripler-directed photoadduct formation at pH 7.0 between the psoralen-linked oligonucleotide and a 30mer duplex represe nting the aromatase target, Photoadduct formation was found to be sens itive to pH, temperature, cation concentration and the base compositio n of the third strand, By varying the base sequence of the target dupl ex around the psoralen intercalation site, we have characterised the s ite and mode of psoralen intercalation, The attached psoralen has been found to intercalate at the tripler-duplex junction with a strong pre ference for one orientation, We have shown that the psoralen will bind at the junction even when there is a preferred TpA step at an adjacen t site, We have also compared the binding affinity and photoreactivity of oligodeoxyribonucleotides linked to two different psoralen derivat ives and found differences in the rate of crosslinking and the extent of crosslink formation, Finally, we have examined oligodeoxyribonucleo tides which are attached to psoralen by polymethylene linkers of diffe rent lengths.