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, 1H-13C-13C-1H total correlation spectroscopy, and 15N-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 1H, 15N, and 13C resonances. Five-hundred eighty-three NOE constraints identified in 3D 13C- and 15N-separated NOE spectroscopy (NOESY)-heteronuclear multiple quantum correlation spectra and a 4-dimensional 13C/13C-edited NOESY spectrum, along with 35 phi, 9 chi 1, and 30 hydrogen bond constraints, were used to calculate 30 structures by hybrid distance geometry/stimulated 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 A 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 A. Several differences were identified some of which may be due to protein-protein interactions in the crystal.
Damberger
F F
FF
Biophysics Graduate Group, University of California, Berkeley 94720.
Pelton
J G
JG
Harrison
C J
CJ
Nelson
H C
HC
Wemmer
D E
DE
D000595:Amino Acid Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry*; D004926:Escherichia coli; D006360:Heat-Shock Proteins; D006860:Hydrogen Bonding; D007716:Kluyveromyces / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D015394:Molecular Structure; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D029701:Saccharomyces cerevisiae Proteins; D012996:Solutions; D014157:Transcription Factors / Q000737:chemistry*
D004268:DNA-Binding Proteins; C067168:HSF1 protein, S cerevisiae; D006360:Heat-Shock Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D012996:Solutions; D014157:Transcription Factors; D004247:DNA