The SKN-1 Amino-Terminal Arm Is a DNA Specificity Segment

The Caenorhabditis elegans SKN-1 protein binds DNA through a basic region like those of bZIP proteins and through a flexible amino-terminal arm segment similar to those with which numerous helix-turn-helix proteins bind to bases in the minor groove. A recent X-ray crystallographic structure suggests that the SKN-1 amino-terminal arm provides only nonspecific DNA binding. In this study, however, we demonstrate that this segment mediates recognition of an AT-rich element that is part of the preferred SKN-1 binding site and thereby significantly increases the sequence specificity with which SKN-1 binds DNA. Mutagenesis experiments show that multiple amino acid residues within the arm are involved in binding. These residues provide binding affinity through distinct but partially redundant interactions and enhance specificity by discriminating against alternate sites. The AT-rich element minor groove is important for binding of the arm, which appears to affect DNA conformation in this region. This conformational effect does not seem to involve DNA bending, however, because the arm does not appear to affect a modest DNA bend that is induced by SKN-1. The data illustrate an example of how a small, flexible protein segment can make an important contribution to DNA binding specificity through multiple interactions and mechanisms.