Post Doctoral Fellow

Biosciences

Johns Hopkins University (Baltimore, MD)

Peter L Privalov

Biophysical examination of bZIP DNA binding proteins.

The bZIP ATF2/cJun transcription factor is a critical component of the human IFN-β enhanceosome. It is widely accepted that the positioning of this heterodimer on the enhancer DNA is determined by other components of this complex, particularly IRF-3 protein. The main purpose of this work was the detailed investigation of the ATF2/cJun association with the enhancer DNA. By the fluorescence anisotropy titration of the FAM and TAMRA labeled fragments of IFN-β DNA it has been shown that the IFN-β enhancer binds at least six ATF2/cJun bZIPs but only one of these heterodimers is bound specifically to the asymmetric PRDIV site with very high affinity, (Kd=3 ± 2 nM). The anisotropy fluorescence titration of the FAM and TAMRA labeled 15 bp DNA duplex containing PRDIV site with the homo and hetero dimers of ATF2 and cJun has been shown that the ATF2/cJun heterodimer binds the asymmetric PRDIV site in definite orientation: the DNA recognition basic segment of ATF2 binds the upstream half-site of the PRDIV site. Thus, the correct positioning of the ATF2/cJun hetero-dimer on the enhancer does not require presence of other components of enhanceosome.