title: Photoaffinity labeling of human retinoid X receptor beta (RXRbeta) with 9-cis-retinoic acid: identification of phytanic acid, docosahexaenoic acid, and lithocholic acid as ligands for RXRbeta.
abstract: We utilized [20-methyl-(3)H]-9-cis-retinoic acid ([(3)H]9-cis-RA) as a direct photoaffinity probe for the characterization of human recombinant retinoid X receptor beta protein (RXRbeta). The photoaffinity labeling was light- and concentration-dependent, saturable, and protected by unlabeled 9-cis-RA in a concentration-dependent manner, indicating that binding occurred in the RXR retinoid binding site. all-trans-Retinoic acid (atRA) did not affect labeling with the 9-cis derivative, confirming that atRA does not compete for the 9-cis-RA binding site. Several retinoid, fatty acid, and bile acid ligands were evaluated for their ability to recognize the 9-cis-RA binding site. Retinol, atRA glucuronide, 13-cis-RA, dolichol, 5,6-epoxy-RA, and vitamin D(3) did not compete for the 9-cis-RA binding site. However, the saturated diterpenoid phytanic acid (PA) and docosahexaenoic acid, which have been recently shown to activate the nuclear receptor, RXR, competed with 9-cis-RA labeling, showing high affinity for the 9-cis-RA binding site. Oleic acid, arachidonic acid, and butyric acid did not interact. However, the bile acid lithocholic acid competed efficiently with 9-cis-RA for the binding site. These data validated the photoaffinity assay as an excellent system for the identification and evaluation of ligands for RXR.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA. RadominskaAnna@exchange.uams.edu
authors.forename: Anna
authors.initials: A
authors.lastname: Radominska-Pandya
authors.forename: Guangping
authors.initials: G
authors.lastname: Chen
mesh_terms: D000077556:Alitretinoin; D001665:Binding Sites; D001667:Binding, Competitive; D004281:Docosahexaenoic Acids / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008024:Ligands; D008095:Lithocholic Acid / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D019920:Photoaffinity Labels; D010831:Phytanic Acid / Q000378:metabolism*; D018168:Receptors, Retinoic Acid / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D047488:Retinoid X Receptors; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D014212:Tretinoin / Q000378:metabolism*
chemical_list: D008024:Ligands; D010446:Peptide Fragments; D019920:Photoaffinity Labels; D018168:Receptors, Retinoic Acid; D011994:Recombinant Proteins; D047488:Retinoid X Receptors; D014157:Transcription Factors; D010831:Phytanic Acid; D000077556:Alitretinoin; D004281:Docosahexaenoic Acids; D014212:Tretinoin; D008095:Lithocholic Acid