Recognition of a 5'-(A,T)GGG(A,T)(2)-3' sequence in the minor groove of DNA by an eight-ring hairpin polyamide

The use of pyrrole-imidazole polyamides for the recognition of core 5'-GGG-3' sequences in the minor groove of double stranded DNA is described. Two hairpin pyrrole-imidazole polyamides, ImImIm-gamma-PyPyPy-beta-Dp and ImImImPy-gamma-PyPyPyPy-beta-Dp (Im = N-methylimidazole-2-carboxamide, Py = N-methylpyrrole-2-carboxamide, beta = p-alanine, gamma = gamma-aminobutyric acid, and Dp = ((dimethylamino)propyl)amide), as well as the corresponding EDTA affinity cleaving derivatives, were synthesized and their DNA binding properties analyzed. Quantitative DNase I footprint titrations demonstrate that ImImIm-gamma-PyPyPy-beta-Dp binds the formal match sequence 5'-AGGGA-3' with an equilibrium association constant of K-a = 5 x 10(6) M(-1) (10 mM Tris . HCl, 10 mM KCl, 10 mM MgCl2, and 5 mM CaCl2, pH 7.0 and 22 degrees C). ImImImPy-gamma-PyPyPyPy-beta-Dp binds the same site, 5'-AGGGAA-3', approximately two orders of magnitude more tightly than the six ring polyamide, with an equilibrium association constant of K-a = 4 x 10(8) M(-1). The eight-ring hairpin polyamide demonstrates greater specifity for single base pair mismatches than does the six ring hairpin. Polyamides with an EDTA . Fe(II) moiety at the carboxy terminus confirm that each hairpin binds in a single orientation. The high affinity recognition of a 5'-GGG-3' core sequence by an eight ring polyamide containing three contiguous imidazole amino acids demonstrates the versatility of pyrrole-imidazole polyamides and broadens the sequence repertoire for DNA recognition.