Stabilization of triple helical DNA by a benzopynidoquinoxaline intercalator

Biophysical, footprinting, and chemical probing experiments are described which characterize the triple helix-stabilizing effects of a benzo[f]pyridoquinoxaline derivative BfPQ-4,3 structurally related to the previously reported benzo[e]pyridoindole compound BePI [Mergny et al. (1992) Science 256, 1681-1684]. Two parallel triple helix model systems have been investigated: one in which the third strand matched perfectly a 27 base pair purine-pyrimidine motif in the target DNA and another in which the third strand was one nucleotide longer, i.e., a 28-mer. In the latter system, the pairing of the (Y)(28) third strand to the (Y . R)(27) target induces the formation of a bulge containing at least one unpaired base, which can be evidenced by chemical probing experiments with osmium tetroxide. BPQ, which unwinds a duplex DNA by 17 degrees as judged by viscometric experiments and otherwise behaves as a typical nonspecific intercalating drug, promotes the formation of a Y . R . Y parallel triple helix containing both T . A . T and C . G . C+ triplets. Both DNase I and MPE . Fe-II footprinting experiments concur that tripler formation with the target (Y . R)(27) sequence can be detected in the presence of BPQ at about 10-fold lower oligonucleotide concentrations than are required to produce an equivalent footprint in the absence of the drug. In addition, BPQ will promote binding to the polypurine-polypyrimidine target sequence by the longer mismatched oligonucleotide, providing significant stabilization of the parallel bulge-containing (Y . R)(27).(Y)(28) triplex with nearly the same efficiency as the bulge-free (Y . R)(27).(Y)(27) tripler. Thus in vivo BPQ might enhance the formation of both undesired and desired DNA triplexes. By performing an MPE . Fe-II probing reaction with a 5'-P-32-labeled oligonucleotide third strand, we have obtained evidence that BPQ is actually bound to the tripler region and may distort it in a sequence-specific fashion.