STRUCTURAL MOTIF OF THE GCN4 DNA-BINDING DOMAIN CHARACTERIZED BY AFFINITY CLEAVING

The NH2-terminal locations of a dimer containing the DNA binding domain of the yeast transcriptional activator GCN4 have been mapped on the binding sites 5′-CTGACTAAT-3′ and 5′-ATGACTCTT-3′. Affinity cleaving was effected by synthetic GCN4 proteins with Fe·EDTA moieties at the NH2-terminus. Analysis of the DNA cleavage patterns for dimers of the Fe·EDTA-proteins corresponding to GCN4 residues 222 to 281 and 226 to 281 revealed that the NH2-termini were in the major groove nine to ten base pairs apart and were symmetrically displaced four to five base pairs from the central C of the recognition site. This result is consistent with the Y-shaped scissor grip-leucine zipper model recently proposed for a class of DNA binding proteins important in the regulation of gene expression.