Base pairing and steric interactions between pyrimidine strand bridging loops and the purine strand in DNA pyrimidine.purine.pyrimidine triplexes

Bimolecular triple-helical DNA complexes recently have found use in a new strategy for the recognition of single-stranded nucleic acids, in which circular (Kool, 1991; Prakash & Kool, 1992) or hairpin-shaped (Giovannangeli et al., 1991; D'Souza & Kool, 1992) oligonucleotides bind these single strands by triplex formation. Bimolecular triplexes may also be formed in vivo as H-DNA, where this structure may potentially play a role in gene expression and recombination (Belotserkovskii et al., 1990; Hanvey et al., 1989; Shimizu et al., 1989). In all of these complexes, the central strand of the triplex must pass beyond the loop that bridges the outer two strands, and models and preliminary experiments have indicated that there may be important interactions between this central strand and the loop (Prakash & Kool, 1992). We now report thermal denaturation studies carried out specifically to investigate these interactions in detail, using as a model the 5'-loop and 3'-loop complexes formed between 14 pyrimidine oligodeoxynucleotides having the sequence 5'-dTTCTTTTCL1TTTL5CTTTTCTT, where L1 and L5 represent varied nucleotides in the loop (which is underlined), and eight target strands having the sequence 5'-dCCCCFAAGAAAAG-3' or 5'-dGAAAAGAAFCCCCC-3', where F is a varied nucleotide flanking the triplex in the central strand. Results correlated from 64 different sequence combinations show that there is wide variation in the stabilities of the complexes, indicating specific and substantial interactions between the nucleotides at the L1, F, and L5 positions. Melting temperatures at pH 7.0 range from 17.0 degrees C to 34.6 degrees C, and free energies (37 degrees C) range from -3.2 to -7.8 kcal mol-1.(ABSTRACT TRUNCATED AT 250 WORDS)