Binding affinities of synthetic peptides, pyridine-2-carboxamidonetropsin and 1-methylimidazole-2-carboxamidonetropsin, that form 2:1 complexes in the minor groove of double-helical DNA.

The designed peptides pyridine-2-carboxamidonetropsin (2-PyN) and 1-methylimidazole-2-carboxamidonetropsin (2-ImN) are crescent-shaped analogs of the natural products netropsin and distamycin A. 2-PyN and 2-ImN bind the 5'-TGTCA-3' sequence as antiparallel side-by-side dimers in the minor groove of DNA. The binding affinities of 2-PyN and 2-ImN to four different 5-bp sites on DNA were determined by quantitative MPE-Fe(II) footprint titration and compared with the tripeptide D from distamycin. The binding affinities of D to the sites 5'-TTTTT-3' and 5'-TGTCA-3' are 2.6 x 10(7) and < 1 x 10(5) M-1, respectively (pH 7.0, 100 mM NaCl). 2-PyN binds these sites with similar affinities, 2.3 x 10(5) and 2.7 x 10(5) M-1, respectively. The affinities of 2-ImN to the same two sites are < 5 x 10(4) and 1.4 x 10(5) M-1, respectively. Substitution of an N-methylpyrrole-2-carboxamide of the distamycin tripeptide by 1-methylimidazole-2-carboxamide has changed the specificities for the two binding sites by a factor of 10(3). The data for 2-PyN and 2-ImN binding the 5'-TGTCA-3' site are best fit by a cooperative binding curve consistent with 2:1 peptide-DNA complexes.