NMR CHARACTERIZATION OF A HETEROCOMPLEX FORMED BY DISTAMYCIN AND ITS ANALOG 2-IMD WITH D(CGCAAGTTGGC)-D(GCCAACTTGCG) - PREFERENCE FOR THE 11/1 2-IMD-DST-DNA COMPLEX OVER THE 2/1 2-IMD-DNA AND THE 2/1 DST-DNA COMPLEXES/

The minor-groove binder distamycin (Dst), its pyrrole-imidazole-pyrrol e analog, 2-imidazole-distamycin (2-ImD), and the oligonucleotide d(CG CAAGTTGGC):d(GCCAACTTGCG) were found to form a 1:1:1 2-ImD-Dst:DNA com plex. As characterized by 2D NOE spectroscopy combined with molecular modeling, one 2-ImD and one Dst molecule bind simultaneously in a head -to-tail orientation contacting the minor groove of the central AAGTT: AACTT site. The 2-ImD ligand lies along the AAGTT strand with the imi dazole nitrogen of the ligand specifically interacting with the guanin e amino group. The distamycin ligand lies along the AACTT strand. The molecular structure of the 1:1:1 2-ImD:Dst:DNA complex is very similar to those of the complexes formed between two 2-ImD molecules (2:1 2-I mD:DNA complex) or two distamycin molecules (2:1 Dst:DNA complex) and the same oligonucleotide duplex. Competition titrations confirm that t he 1:1:1 2-ImD:Dst:DNA complex forms preferentially over the 2:1 2-ImD :DNA complex, as well as over the newly discovered 2:1 Dst:DNA complex . These results indicate that (i) the hydrogen-bond-accepting imidazol e nitrogen of the 2-ImD ligand in the 1:1:1 2-ImD:Dst:DNA complex is r esponsible for the strand-specific recognition of a GC base pair, (ii) the availability of a single hydrogen bond acceptor on the ligand mol ecule per guanine amino group enhanced both specificity and affinity o f DNA binding, and (iii) different distamycin-like ligands can be comb ined in the 2:1 binding motif to expand the range of DNA sequences tha t can be specifically recognized through the minor groove.