Participation of the amino-terminal domain in the self-association of the full-length yeast TATA binding protein

The association of monomeric TATA binding protein with promoter DNA is an e ssential first step in many current models of eukaryotic transcription init iation. This step is followed by others in which additional transcription f actors, and finally RNA polymerase, assemble at the promoter. Here we chara cterize the quaternary interactions of the Saccharomyces cerevisiae TATA-bi nding protein (yTBP), in the absence of other proteins or DNA. The data rev eal a robust pattern in which yTBP monomers equilibrate with tetramers and octamers over a broad span of temperatures (4 degrees C less than or equal to T less than or equal to 37 degrees C) and salt concentrations (60 mM les s than or equal to [KCl] less than or equal to 1 M), that includes the phys iological range. Association is highly cooperative, with octamer formation favored by similar to 9 kcal/mol over tetramer formation. Changes in associ ation constant with [KCl] are consistent with an assembly-linked release of ions at low salt and an assembly-linked uptake of ions at high salt, for b oth monomer reversible arrow tetramer and tetramer reversible arrow octamer reaction steps. Fluorescence emission spectra and steady-state anisotropie s reveal that the amino-terminal domain changes conformation and dynamics a t both association steps and that the polarity of the environment near tryp tophan 26 is sensitive to changes in [KCl] in the monomeric and tetrameric states but not the octameric state. These results are consistent with a [sa lt]-dependent change in the assembly mechanism near 300 mM KCl and suggest that the amino-terminal domain may modulate the self-association of the ful l-length protein. TBP self-association may regulate many of its cellular fu nctions, including transit of the nuclear membrane and participation in tra nscription initiation.