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DNA-Binding Oligoamides

DNA-binding oligoamides are heterocyclic crescent shaped molecules composed of pyrrole and imidazole-based rings [96-98]. Three different heterocyclic rings are commonly used: N-methylpyrrole (Py), N-methylimidazole (Im), and N-methyl-3-hydroxypyrrole (Hp) (Fig. 8.2). These oligoamides were initially designed based on the natural product Distamycin A, which binds A/T-rich se-

lm Hp Py

Fig. 8.2 Chemical structures of Distamycin A and pyrole and imidazole based monomers of DNA-binding olioamides (Im = N-methylimidazole, Hp = N-methyl-3-hydroxypyrrole, Py = N-methylpyrrole).

lm Hp Py

Fig. 8.2 Chemical structures of Distamycin A and pyrole and imidazole based monomers of DNA-binding olioamides (Im = N-methylimidazole, Hp = N-methyl-3-hydroxypyrrole, Py = N-methylpyrrole).

quences in the DNA minor groove as 1:1 or 2:1 Distamycin A-DNA complexes [99]. These oligoamides have been used as site-specific DNA-targeting agents with affinities comparable to DNA-binding proteins [97]. These DNA-binding oligoamides do not adopt specific conformations unless bound to DNA, and therefore are considered foldamers in a liberal sense. Crystal structures revealed that the ring amide groups of these oligoamides recognize the edges of adjacent oligonucleotide strands through a series of hydrogen bonds [100-102]. ''Pairing rules'' for targeting specific sequences of DNA with oligoamide base pairs have been deduced [100-103]. Py/Hp recognizes A-T; Hp/Py recognizes T-A; Py/Im recognizes C-G; Im/Py recognizes G-C.

Structural data of oligoamide-DNA complexes show that the oligoamide rise per residue matches the pitch of the B-form DNA [100-102]. However, the curvature of the oligoamide crescent shape is slightly greater than that of the minor groove of DNA, thus oligoamides longer than 5 units do not completely complement the target DNA, limiting the size of DNA that can be targeted [100]. However, binding sites 10-11 base pairs long have been targeted by oligoamides containing curvature-relaxing moieties such as b-Ala [104] or by multiple oligoamides linked to form hairpin structures [105-108].

Oligoamides can be synthesized through solid phase methods employing Boc-or Fmoc-protected monomers [109-111]. The first residue attached to the resin determines the oligoamide C-terminal residue, which can dictate the DNA-binding specificity. For example, oligoamides with a C-terminal b-Ala specifically target T-A pairs [112] whereas oligoamides without the C-terminal b-Ala target G-C pairs [113].

Localization to nuclei is crucial for DNA targeting with oligoamides. Nuclear localization of oligoamides depends on numerous factors, including the cell type being targeted, oligoamide characteristics (such as charge, shape, sequence), and the identity and placement of dye conjugates [114-117]. Idiosyncratic determinants of nuclear localization have also been observed; the presence of a C-terminal b-Ala residue or the absence of an alkyl amine moiety inhibited nuclear localization, while the presence of an acetylated 2,4-diaminobutyric acid turn promotes nuclear localization [117]. However, nuclear exclusion and localization to other cellular compartments (i.e. lysosomes) often occurs [114, 115].

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