THE ACTIVATING TRANSCRIPTION FACTOR REGION WITHIN THE E2A PROMOTER EXISTS IN A NOVEL CONFORMATION

The ATF (activating transcription factor) binding site within the E2A promoter region of adenovirus is shown to exist in a novel conformation in vitro via nuclear magnetic resonance methods. This novel conformation may be important to the protein DNA recognition process. This conformation has characteristics of both A- and B-form DNA. From circular dichroism and through-space-based NMR experiments, it is clear that the overall helical structure is B-like. However, the H-1-H-1 coupling constant information indicates that most of the sugar puckers of the individual nucleotides are in the C3'-endo/C4'-exo range which is more characteristic of A-form DNA. The sugar conformation can also be described as a mixture of two states, C3'-endo and C2'-endo, where many of the sugars exist mainly in the C3'-endo state. These data show that the conformation of the sugar puckers does not determine the nature of the overall base stacking on the DNA. Helical parameters were calculated from NOE build-up curves for half of the dinucleotide pairs. Severe spectral overlap on the nuclear Overhauser based spectra prevented determination of the helical parameters for all of the dinucleotide base-pairs. Energy minimization and molecular dynamic simulation methods using the sugar pucker and glycosidic torsion angles determined from the NMR data as constraints were carried out in order to demonstrate that such a conformation was energetically favorable.