Deconstruction of GCN4/GCRE into a monomeric peptide-DNA complex

Here we describe a system that enables short peptides to bind DNA sequence-specifically. Linking the peptide covalently to DNA through a disulphide bond eliminates the unfavourable energetic cost of diffusion and thus potentiates the peptide-DNA interaction. By this approach we have deconstructed the GCN4/DNA complex into its elemental DNA recognition units. We find that the GCN4 basic region contacts the two half-sites with very different affinities and propose that this thermodynamic asymmetry plays a role in differential regulation of gene expression. Specific binding of the peptide to DNA stabilizes the disulphide bond toward reduction suggesting a novel approach to the discovery of new DNA-binding specificities.