Signals for TBP/TATA box recognition.
The TATA box-binding protein (TBP) recognizes its target sites (TATA boxes) by indirectly reading the DNA sequence through its conformation effects (indirect readout). Here, we explore the molecular mechanisms underlying indirect readout of TATA boxes by TBP by studying the binding of TBP to adenovirus major late promoter (AdMLP) sequence variants, including alterations inside as well as in the sequences flanking the TATA box. We measure here the dissociation kinetics of complexes of TBP with AdMLP targets and, by phase-sensitive assay, the intrinsic bending in the TATA box sequences as well as the bending of the same sequence induced by TBP binding. In these experiments we observe a correlation of the kinetic stability to sequence changes within the TATA recognition elements. Comparison of the kinetic data with structural properties of TATA boxes in known crystalline TBP/TATA box complexes reveals several "signals" for TATA box recognition, which are both on the single base-pair level, as well as larger DNA tracts within the TATA recognition element. The DNA bending induced by TBP on its binding sites is not correlated to the stability of TBP/TATA box complexes. Moreover, we observe a significant influence on the kinetic stability of alteration in the region flanking the TATA box. This effect is limited however to target sites with alternating TA sequences, whereas the AdMLP target, containing an A tract, is not influenced by these changes.
Bareket-Samish
A
A
Department of Biology, Technion, Technion City, Haifa, 32000, Israel. bitali@tx.technion.ac.il
Cohen
I
I
Haran
T E
TE
D000256:Adenoviridae / Q000235:genetics; D020029:Base Pairing / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D016384:Consensus Sequence / Q000235:genetics; D004247:DNA / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006207:Half-Life; D007700:Kinetics; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation; D018583:Pliability; D011485:Protein Binding; D013379:Substrate Specificity; D016385:TATA Box / Q000235:genetics*; D035181:TATA-Box Binding Protein; D013816:Thermodynamics; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D015003:Yeasts
D004268:DNA-Binding Proteins; D035181:TATA-Box Binding Protein; D014157:Transcription Factors; D004247:DNA