NMR chemical shift perturbation mapping of DNA binding by a zinc-finger domain from the yeast transcription factor ADR1

Mutagenesis studies have revealed that the minimal DNA-binding domain of the yeast transcription factor ADR1 consists of two CYS2-His(2) zinc fingers plus an additional 20 residues proximal and N-terminal to the fingers. We have assigned NMR H-1, N-15, and C-13 chemical shifts for the entire minimal DNA-binding domain of ADR1 both free and bound to specific DNA. H-1 chemical shift values suggest little structural difference between the zinc fingers in this construct and in single-finger constructs, and C-13(alpha) chemical shift index analysis indicates little change in finger structure upon DNA binding. H-1 chemical shift perturbations upon DNA binding are observed, however, and these are mapped to define the protein-DNA interface. The two zinc fingers appear to bind DNA with different orientations, as the entire helix of finger I is perturbed, while only the extreme N-terminus of the finger 2 helix is affected. Furthermore, residues N-terminal to the first finger undergo large chemical shift changes upon DNA binding suggesting a role at the protein-DNA interface. A striking correspondence is observed between the protein-DNA interface mapped by chemical shift changes and that previously mapped by mutagenesis.