Sequence specific alkylation of DNA by hairpin pyrrole–imidazole polyamide conjugates Original Research Article

Background: Pyrrole–imidazole polyamides are synthetic ligands that recognize predetermined sequences in the minor groove of DNA with affinities and specificities comparable to those of DNA-binding proteins. As a result of their DNA-binding properties, polyamides could deliver reactive moieties for covalent reaction at specific DNA sequences and thereby inhibit DNA–protein interactions. Site-specific alkylation of DNA could be a useful tool for regulating gene expression. As a minimal first step, we set out to design and synthesize a class of hairpin polyamides equipped with DNA alkylating agents and characterize the specificity and yield of covalent modification. Results: Bis(dichloroethylamino)benzene derivatives of the well-characterized chlorambucil (CHL) were attached to the γ turn of an eight-ring hairpin polyamide targeted to the HIV-1 promoter. We found that a hairpin polyamide–CHL conjugate binds and selectively alkylates predetermined sites in the HIV promoter at subnanomolar concentrations. Cleavage sites were determined on both strands of a restriction fragment containing the HIV-1 promoter, revealing good specificity and a high yield of alkylation. Conclusions: The ability of polyamide–CHL conjugates to sequence specifically alkylate double-stranded DNA in high yield and at low concentrations sets the stage for testing their use as regulators of gene expression in cell culture and ultimately in complex organisms. Article Outline