INHIBITION OF MAJOR-GROOVE-BINDING PROTEINS BY PYRROLE-IMIDAZOLE POLYAMIDES WITH AN ARG-PRO-ARG POSITIVE PATCH

Background: Gene-specific targeting of any protein-DNA complex by smal l molecules is a challenging goal at the interface of chemistry and bi ology. Polyamides containing N-methylimidazole and N-methylpyrrole ami no acids are synthetic ligands that have an affinity and specificity f or DNA comparable to many naturally occurring DNA-binding proteins. It has been shown that an eight-ring hairpin polyamide targeted to a spe cific minor-groove contact within a transcription factor binding site can inhibit protein-DNA binding and gene transcription. Polyamides and certain major-groove-binding proteins have been found to cc-occupy th e DNA helix, however. To expand the number of genes that can be target ed by pyrrole/imidazole polyamides, we set out to develop a class of p olyamides that can selectively inhibit major-groove-binding proteins. Results: An eight-ring hairpin polyamide conjugated to a carboxy-termi nal Arg-Pro-Arg tripeptide was designed to deliver a positive residue to the DNA backbone and interfere with protein-phosphate contacts. Gel mobility shift analysis demonstrated that a polyamide hairpin-Arg-Pro -Arg binding in the minor groove selectively inhibits binding of the t ranscription factor GCN4 (222-281) in the adjacent major groove. Subst itution within the Arg-Pro-Arg revealed that each residue was required for optimal GCN4 inhibition. Conclusions: A pyrrole-imidazole polyami de that binds to a predetermined site in the DNA minor groove and deli vers a positive patch to the DNA backbone can selectively inhibit a DN A-binding protein that recognizes the adjacent major groove. A subtle alteration of the DNA microenvironment targeted to a precise location within a specific DNA sequence could achieve both gene-specific and pr otein-specific targeting.