Characterization of a Max:DNA Complex by Cross-Linking to Photoactive Oligonucleotides

Abstract

The structure of the Max:DNA complex was investigated by cross-linking Max to a series of photoactive oligonucleotides. A single photoactive aryl azide was introduced into oligonucleotides at defined positions downstream from the specific CACGTG binding site. Purified Max homodimers bound to and were cross-linked to oligonucleotides containing a photoactive group either 2, 5, 8, or 11 bp downstream from the binding site. Further analysis revealed that an amino-terminal chymotryptic peptide of Max was cross-linked to the oligonucleotide containing a photoactive probe 11 bp downstream from the specific binding site. This result is consistent with the recent crystal structure of the Max:DNA complex (Ferré-D'Amaré et al., 1993) and further suggests that amino acid residues near the amino-terminus of Max are in close proximity to a region of DNA that is separated from the core binding site by one turn of the double helix.