A series of mutations in the homeo domain of the yeast α2 protein were constructed to test, both in vivo and in vitro, predictions based on the α2- DNA cocrystal structure described by Wolberger et al. (1991). The effects of the mutations were observed in three different contexts using authentic target DNA sequences: α2 binding alone to specific DNA, α2 binding cooperatively with MCM1 to specific DNA, and α2 binding cooperatively with al to specific DNA. As expected, changes in the amino acid residues that contact DNA in the X-ray structure severely compromised the ability of α2 to bind DNA alone and to bind DNA cooperatively with MCM1. In contrast, many of these same mutations, including a triple change that altered all the 'recognition' residues of helix 3, had little or no effect on the cooperative binding of α2 and a1 to specific DNA, as determined both in vivo and in vitro. These results show that the ability of a homeo domain protein to correctly select and repress target genes does not necessarily depend on the residues commonly implicated in sequence-specific DNA binding.
All Science Journal Classification (ASJC) codes
- Developmental Biology