Structural and thermodynamic characterization of the DNA binding properties of a triple alanine mutant of MATα2

Triply mutated MATα2 protein, α2-3A, in which all three major groove-contacting residues are mutated to alanine, is defective in binding DNA alone or in complex with Mcm1 yet binds with MATa1 with near wild-type affinity and specificity. To gain insight into this unexpected behavior, we determined the crystal structure of the a1/α2-3A/DNA complex. The structure shows that the triple mutation causes a collapse of the α2-3A/DNA interface that results in a reorganized set of α2-3A/DNA contacts, thereby enabling the mutant protein to recognize the wild-type DNA sequence. Isothermal titration calorimetry measurements reveal that a much more favorable entropic component stabilizes the a1/α2-3A/DNA complex than the α2-3A/DNA complex. The combined structural and thermodynamic studies provide an explanation of how partner proteins influence the sequence specificity of a DNA binding protein.