STEREOCHEMICAL CONTROL OF THE DNA-BINDING AFFINITY, SEQUENCE SPECIFICITY, AND ORIENTATION PREFERENCE OF CHIRAL HAIRPIN POLYAMIDES IN THE MINOR-GROOVE

Three-ring polyamides containing pyrrole (Py) and imidazole (Im) amino acids covalently coupled by gamma-aminobutyric acid (gamma) form six- ring hairpins that recognize five-base-pair sequences in the minor gro ove of DNA. Selective chiral substitution of the ''gamma-turn'' enhanc es the properties of polyamide hairpins with regard to DNA affinity an d sequence specificity. Polyamides of core sequence composition ImPyPy -gamma-PyPyPy which differ by selective stereochemical substitution of the prochiral alpha-position in the gamma-turn were prepared. The DNA binding properties of two enantiomeric polyamides were analyzed by fo otprinting and affinity cleavage on a DNA. fragment containing two mat ch sites (5'-TGTTA-3' and 5'-ACATT-3') and one 5'-TGTCA-3' mismatch si te. Quantitative footprint titrations demonstrate that replacement of gamma-aminobutyric acid by (R)-2,4-diaminobutyric acid enhances DNA bi nding affinity for the 5'-TGTTA-3' match site 13-fold (K-a = 3.8 x 10( 9) M-1). The enhanced affinity is achieved without a compromise in seq uence selectivity, which in fact increases and is found to be 100-fold higher relative to binding at a single base pair mismatch sequence, 5 '-TGTCA-3'. An (S)-2,4-diaminobutyric acid linked hairpin binds with 1 70-fold reduced affinity relative to the R-enantiomer and only 5-fold sequence specificity versus a 5'-ACATT-3' reversed orientation site. T hese effects are modulated by acetylation of thr chiral amine substitu ents. This study identifies structural elements which should facilitat e the design of new hairpin polyamides with improved DNA binding affin ity, sequence specificity, and orientational selectivity.