SOLUTION STRUCTURE OF THE DNA-BINDING DOMAIN OF THE HEAT-SHOCK TRANSCRIPTION FACTOR DETERMINED BY MULTIDIMENSIONAL HETERONUCLEAR MAGNETIC-RESONANCE SPECTROSCOPY

The solution structure of the 92-residue DNA-binding domain of the heat shock transcription factor from Kluyveromyces lactis has been determined using multidimensional NMR methods. Three-dimensional (3D) triple resonance, H-1-C-13-C-13-H-1 total correlation spectroscopy, and N-15-separated total correlation spectroscopy-heteronuclear multiple quantum correlation experiments were used along with various 2D spectra to make nearly complete assignments for the backbone and side-chain H-1, N-15, and C-13 resonances. Five-hundred eighty-three NOE constraints identified in 3D C-13- and N-15-separated NOE spectroscopy (NOESY)-heteronuclear multiple quantum correlation spectra and a 4-dimensional C-13/C-13-edited NOESY spectrum, along with 35 phi, 9(chi 1), and 30 hydrogen bond constraints, were used to calculate 30 structures by a hybrid distance geometry/simulated annealing protocol, of which 24 were used for structural comparison. The calculations revealed that a 3-helix bundle packs against a small 4-stranded antiparallel beta-sheet. The backbone RMS deviation (RMSD) for the family of structures was 1.03 +/- 0.19 Angstrom with respect to the average structure. The topology is analogous to that of the C-terminal domain of the catabolite gene activator protein and appears to be in the helix-turn-helix family of DNA-binding proteins. The overall fold determined by the NMR data is consistent with recent crystallographic work on this domain (Harrison CJ, Bohm AA, Nelson HCM, 1994, Science 263:224) as evidenced by RMSD between backbone atoms in the NMR and X-ray structures of 1.77 +/- 0.20 Angstrom. Several differences were identified some of which may be due to protein-protein interactions in the crystal.