AFFINITY, STABILITY AND POLARITY OF BINDING OF THE TATA-BINDING PROTEIN GOVERNED BY FLEXURE AT THE TATA BOX

The TATA binding protein (TBP), which plays a central role in gene reg ulation as an essential component of all three nuclear transcription s ystems, sharply kinks the TATA box at two sites and severely contorts the intervening DNA segment. DNA constructs with precisely localized f lexure have been used to investigate the special repertoire of mechani sms and properties that arise from TBP interacting with the TATA box. DNA flexure precisely localized to the sites of TBP-mediated DNA kinki ng increases the affinity of TBP more than 100-fold; unexpectedly, thi s increase in affinity is achieved almost exclusively by increasing th e stability of the TBP-DNA complex rather than the rate of its formati on. In vitro transcription with RNA polymerase III provides a first de monstration that the orientation of TBP on the TATA box is governed by DNA deformability, its C-proximal repeat contacting the more flexible end of the TATA box. Exceptionally stable TBP-DNA complexes reach the ir orientational equilibrium very slowly; in these circumstances, asse mbly of stable (''committed'') transcription initiation complexes can freeze far-from-equilibrium orientations of TBP on the TATA box, causi ng transcription polarity to be determined by a kinetic trapping mecha nism. (C) 1998 Academic Press.