Sequence specific alkylation of DNA by hairpin pyrrole-imidazole polyamide conjugates

Abstract

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.

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