EXPRESSION, PURIFICATION, AND FUNCTIONAL-ANALYSIS OF THE DNA-BINDING DOMAIN OF THE NUCLEAR RECEPTOR REV-ERB-BETA

Rev-erb beta is a member of the nuclear receptor superfamily, which in cludes a group of transcription factors involved in the response to st eroids, vitamin D, retinoic acids, and other lipophilic molecules. The Rev-erb nuclear receptors exist at least in two forms, namely Lu and beta, with a high degree of evolutionary conservation at the level of the DNA binding domain. Nevertheless, the exact type of DNA binding of these proteins is not fully understood, In order to get insight into this DNA binding mechanism we obtained a pure and functional homogeneo us recombinant protein in bacteria corresponding to the DNA binding do main of Rev-erb beta (REDBD). REDBD interacts with oligonucleotides co ntaining an A/T-rich sequence preceding a single AGGTCA site (Rev-RE) or with an AGGTCA direct repeat separated by 2 bp (Rev-DR2). These res ults, and the affinity parameters of the interaction between REDBD and Rev-RE or Rev-DRa, indicate that our REDBD preparation is capable of specific and tight binding to DNA targets. (C) 1996 Academic Press, In c.