PROTON AND NITROGEN NMR SEQUENCE-SPECIFIC ASSIGNMENTS AND SECONDARY STRUCTURE DETERMINATION OF THE BACILLUS-SUBTILIS SPO1-ENCODED TRANSCRIPTION FACTOR-1

Sequence-specific H-1 and N-15 NMR(1) assignments are reported for the transcription factor 1 (TF1), a 22-kDa type II DNA-binding protein (D BPII) that consists of two 99-residue monomers. An assignment strategy is employed that uses six complementary selectively deuterium-labeled TF1 variants and an uniformly N-15-labeled TF1 variant. Two-dimension al and three-dimensional homonuclear and heteronuclear NMR correlated spectra are analyzed and yield nearly complete assignments for the H-1 and N-15 resonances. Discrete protein secondary structure domains are also defined; in each monomer, three alpha-helices, an antiparallel b eta-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 intermono meric or intramonomeric. An analysis of amide proton exchange reveals that the carboxyterminal alpha-helix is less stable than the other two alpha-helices in each monomer. A previously proposed working structur al model of the TF1 dimer [Geiduschek et al. (1990) J. Struct. Biol. 1 04, 84-90], based on the crystal structure of a highly homologous DBPI I, the Bacillus stearothermophilus-encoded HU protein, is generally su pported by our results. Several departures from this model, however, a re noted. Most notably, the carboxy-terminal tail of TF1 adopts an alp ha-helical conformation with a backbone distortion at Lys93.