Linked polyamides are currently the most promising compounds for the creation of bioavailable sequence-specific DNA-binding molecules for use in chemotherapy and biosensing (1). These compounds are composed of two linked polyamide chains, analogous to the antibiotics netropsin and distamycin, running side-by-side and antiparallel down a widened minor groove of B-DNA, with a polyamide ring packed tightly against each DNA base, as diagrammed in Fig. 1. Pyrrole-imidazole polyamides, with either a hairpin linkage (2) or a central "stapled" linkage (3), bind in the minor groove of DNA in a partially sequence-specific manner. Pyrrole is the naturally occurring G-excluding element of netropsin and distamycin (4, 5), and the imidazole ring was first proposed as a G-reading element based on the structures of 1:1 polyamide:DNA complexes (6-8). Footprinting analysis has demonstrated the pairing rules for these rings in hairpin-linked polyamides: imidazole-pyrrole pairs bind strongly to GC bp and, by symmetry, pyrrole-imidazole pairs bind to CG, whereas pyrrole-pyrrole pairs are degenerate, binding strongly to both AT and TA (9). NMR (10-12) and crystallographic (13-15) analyses have revealed the specific steric and hydrogen bonding interactions that mediate this specificity.
This report analyzes the theoretical limits of DNA sequence discrimination by linked polyamides composed of two to four different types of rings, each preferentially binding to a different base. An ideal sequence-reading polyamide, or "lexitropsin" (6-8), with full base-reading ability would be built from four different types of rings, each binding specifically to one of the four DNA bases. Unfortunately, such hyper-specific rings have not been discovered and given the close similarity of the minor groove faces of the two pyrimidines, may never be discovered. This report examines the optimal design of polyamides composed of less than this perfect complement of rings, which were chosen to maximize the fraction of polyamide bound to the target DNA sequence. Two design issues are addressed: (i) the optimal choice of base-binding specificity for each ring, and (ii) the optimal polyamide composed of these rings designed to target a given DNA sequence. A full mathematical analysis will be presented in a separate publication; this report presents the major implications for polyamide design.
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