NMR EVIDENCE FOR SIMILARITIES BETWEEN THE DNA-BINDING REGIONS OF DROSOPHILA-MELANOGASTER HEAT-SHOCK FACTOR AND THE HELIX-TURN-HELIX AND HNF-3/FORKHEAD FAMILIES OF TRANSCRIPTION FACTORS

Heteronuclear multidimensional NMR experiments of residues 33-163 of the DNA-binding domain of Drosophila heat shock factor, dHSF(33-163), were recorded, using only 3 mg of uniformly N-15-labeled or 2 mg of uniformly N-15/C-13-labeled protein. The polypeptide consists of a structured part comprising three helices, a three-stranded antiparallel beta-sheet, with the first two strands connected by a four-residue type I tight turn. The second helix is disrupted at its C-terminal end by a proline residue and is followed by an extended turn, leading to the third helix. The dHSF(33-163) protein is unstructured at its N- and C-termini, and a third unstructured region is found from Thr113 to Arg124. Exchange broadening of the N-15-H-1 correlations upon titration of N-15 labeled HSF with a 13-base-pair DNA duplex suggests a DNA-binding motif in which the third helix acts as the recognition helix. Both the secondary structure and DNA-binding pattern of dHSF(33-163) suggest that the overall topology resembles that the helix-turn-helix bacterial activator CAP [Weber, 1. T., & Steitz, T. A. (1987) J. Mol. Biol. 198, 311-326] and the liver-specific transcription factor HNF-3gamma, the prototype of the HNF-3/forkhead protein family [Clark, K. L., Halay, E. D., Lai, E., & Burley, S. K. (1993) Nature 364, 412-420].