Proton and nitrogen NMR sequence-specific assignments and secondary structure determination of the Bacillus subtilis SPO1-encoded transcription factor 1.
Sequence-specific 1H and 15N NMR1 assignments are reported for the transcription factor 1 (TF1), a 22-kDa type II DNA-binding protein (DBPII) that consists of two 99-residue monomers. An assignment strategy is employed that uses six complementary selectively deuterium-labeled TF1 variants and an uniformly 15N-labeled TF1 variant. Two-dimensional and three-dimensional homonuclear and heteronuclear NMR correlated spectra are analyzed and yield nearly complete assignments for the 1H and 15N resonances. Discrete protein secondary structure domains are also defined; in each monomer, three alpha-helices, an antiparallel beta-sheet, and an antiparallel beta-ribbon are identified. Analyses of two dimers formed from two distinct selectively deuteriated monomers serve to identify a number of interproton contacts as either intermonomeric or intramonomeric. An analysis of amide proton exchange reveals that the carboxy-terminal alpha-helix is less stable than the other two alpha-helices in each monomer. A previously proposed working structural model of the TF1 dimer [Geiduschek et al. (1990) J. Struct. Biol. 104, 84-90], based on the crystal structure of a highly homologous DBPII, the Bacillus stearothermophilus-encoded HU protein, is generally supported by our results. Several departures from this model, however, are noted. Most notably, the carboxy-terminal tail of TF1 adopts an alpha-helical conformation with a backbone distortion at Lys93.
Jia
X
X
Department of Chemistry, University of California at San Diego, La Jolla 92093.
Reisman
J M
JM
Hsu
V L
VL
Geiduschek
E P
EP
Parello
J
J
Kearns
D R
DR
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry*; D006860:Hydrogen Bonding; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D014157:Transcription Factors / Q000737:chemistry*; D014764:Viral Proteins
D004268:DNA-Binding Proteins; C059361:TF1 DNA-binding protein, Bacteriophage SPO1; D014157:Transcription Factors; D014764:Viral Proteins