Expanding the recognition of the minor groove of DNA by incorporation of β-alanine in hairpin polyamides

In order to expand the recognition code by hairpin polyamides to include DNA sequences of the type 5'-CWWC-3' two polyamides, PyPyPyPy-(R)(H2N)gamma -ImPyPyIm-beta -Dp (1) and PyPyPyPy-(R)(H2N)gamma -ImPy-beta -Im-beta -Dp (2) were synthesized which have in common an Py/1m pair in the terminal position for targeting CG but differ with respect to internal placement of a beta -alanine residue. The equilibrium association constants (K-a) were determined at four DNA sites which differ at a single common position, 5'-TNTACA-3' (N = T, A, G, C). Quantitative DNase I footprint titration experiments reveal that the eight-ring hairpin PyPyPyPy-(R) (H2N)y-ImPyPyIm-beta -Dp (1) binds the four binding sites with similar affinities, K-a = 1.3-1.9x10(10) M-1 indicating that there is no preference for the position N. In contrast, a redesigned polyamide PyPyPyPy-(R)(H2N)gamma -ImPy-beta -Im-beta -Dp (2) that places an internal flexible aliphatic beta -alanine to the 5'-side of a key imidazole group bound the match site 5'-TCTACA-3' with high affinity and good sequence discrimination (K-a(match) = 4.9 x 10(10) M-1 and the single base pair mismatch sites with 5- to 25-fold lower affinity). These results expand the repertoire of sequences targetable by hairpins and emphasize the importance of beta -alanine as a key element for minor groove recognition. (C) 2001 Elsevier Science Ltd. All rights reserved.