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.
Stanojevic
D
D
Department of Chemistry, Harvard University, Cambridge, Massachusetts 02138, USA.
Verdine
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GL
D000595:Amino Acid Sequence; D001483:Base Sequence; D002942:Circular Dichroism; D016384:Consensus Sequence; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004220:Disulfides / Q000737:chemistry; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016350:Leucine Zippers / Q000502:physiology*; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D013329:Structure-Activity Relationship; D013439:Sulfhydryl Reagents / Q000737:chemistry; D013696:Temperature
D004268:DNA-Binding Proteins; D004220:Disulfides; D005656:Fungal Proteins; D010455:Peptides; D029701:Saccharomyces cerevisiae Proteins; D013439:Sulfhydryl Reagents; D004247:DNA; D011494:Protein Kinases