Structural features of TBP/TATA-box Interaction
The TATA binding protein (TBP) is a key protein in the assembly of eukaryotic transcriptional preinitiation complexes. The TATA box is the TBP target in RNA polymerase II promoters. The recognition of TATA boxes by TBP involves indirect reading of the DNA sequence, which means recognition of the intrinsic structure of the DNA target or its deformation upon complex formation. We studied the binding of TBP to Adenovirus major late promoter (AdMLP) sequence variants, including alterations both inside and in the sequences flanking the TATA box. We measured the dissociation rate of TBP from the AdMLP variants, and determined, by phase-sensitive assay, the intrinsic bend angle of these targets, as well as the bend angles induced on the DNA double helix upon binding to TBP. We show that sequences flanking the core 8-bp TATA box affect the stability of binding of TBP to its target sites, although there are no direct contacts between TBP and these regions. However, this effect is sequence dependent. Sequences flanking TATA boxes, which contain the more rigid A4 tract, are not influenced by the nature of these sequences, whereas they will perturb TATA elements containing alternating (T-A) sequences, which are more mechanically flexible. Moreover, we find that sequence changes withinTATA boxes, which lead to more flexible TATA boxes form less stable complexes with TBP. The mechanical flexibility of certain individual base-pair steps, and especially that of the entire binding site is correlated with kinetic instability of the complexes. We do not observe a correlation between TBP-induced TATA-box bending and the measured stability of these complexes, as been observed in the past, and in other systems. Based on our results we propose structural signals involved in TATA-box recognition, which may suggest how TBP can differentially decode the information encoded in the sequence of different TATA boxes, although the TBP/TATA-box interaction is exclusively through the minor groove.
Tali. E. Haran1, Avital Bareket-Samish2 and Ilana Cohen
Department of Biology, Technion, Technion City, Haifa 32000, Israel. 2present address: Department of Structural Biology, Fairchild Science Building, D-125, Stanford University School of Medicine, Stanford CA 94305-5400 USA. 1e-mail of the corresponding author: firstname.lastname@example.org