title: Characterization of DNA binding and retinoic acid binding properties of retinoic acid receptor.
abstract: High-level expression of the full-length human retinoic acid receptor (RAR) alpha and the DNA binding domain of the RAR in Escherichia coli was achieved by using a T7 RNA polymerase-directed expression system. After induction, full-length RAR protein was produced at an estimated level of 20% of the total bacterial proteins. Both intact RAR molecules and the DNA binding domain bind to the cognate DNA response element with high specificity in the absence of retinoic acid. However, this binding is enhanced to a great extent upon the addition of eukaryotic cell extracts. The factor responsible for this enhancement is heat-sensitive and forms a complex with RAR that binds to DNA and exhibits a distinct migration pattern in the gel-mobility-shift assay. The interaction site of the factor with RAR is localized in the 70-amino acid DNA binding region of RAR. The hormone binding ability of the RAR alpha protein was assayed by a charcoal absorption assay and the RAR protein was found to bind to retinoic acid with a Kd of 2.1 x 10(-10) M.
authors.affiliation: Department of Chemistry, University of California, San Diego, La Jolla 92093.
authors.forename: N
authors.initials: N
authors.lastname: Yang
authors.forename: R
authors.initials: R
authors.lastname: Schüle
authors.forename: D J
authors.initials: DJ
authors.lastname: Mangelsdorf
authors.forename: R M
authors.initials: RM
authors.lastname: Evans
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004331:Drosophila melanogaster; D006367:HeLa Cells; D006801:Humans; D066298:In Vitro Techniques; D008024:Ligands; D009687:Nuclear Proteins / Q000378:metabolism; D011485:Protein Binding; D018168:Receptors, Retinoic Acid; D011994:Recombinant Proteins / Q000378:metabolism; D014158:Transcription, Genetic; D014212:Tretinoin / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D008024:Ligands; D009687:Nuclear Proteins; D018168:Receptors, Retinoic Acid; D011994:Recombinant Proteins; D014212:Tretinoin