title: Helix 1/8 interactions influence the activity of nuclear receptor ligand-binding domains.
abstract: The ligand-binding domain (LBD) of apo-nuclear receptors in solution is thought to be a very dynamic structure with many possible conformations. Upon ligand binding, the structure is stabilized to a more rigid conformation. The dynamic stabilization assay is a LBD reassembly assay that takes advantage of the high specificity of the intramolecular interactions that comprise the ligand-bound LBD. Here, we demonstrate dynamic stabilization for the nuclear receptors peroxisome proliferator-activated receptor (PPAR)gamma and nerve growth factor inducible (NGFIB)beta and identify residues important for stabilization of the intramolecular interactions induced by PPARgamma ligands. Site-directed mutagenesis studies identified residues in helices 1 and 8 required for LBD reassembly. Further, disrupting the helix 1/8 interaction in the context of the holo-LBD alters the response of the receptor in a compound-specific manner, suggesting that residues far from the ligand-binding pocket can influence the stability of the ligand-bound receptor. Thus, these results support and extend models of the apo-LBD of PPARgamma as a dynamic structure.
authors.affiliation: GlaxoSmithKline Research and Development, Research Triangle Park, North Carolina 27709-3398, USA.
authors.forename: Jason A
authors.initials: JA
authors.lastname: Holt
authors.forename: Thomas G
authors.initials: TG
authors.lastname: Consler
authors.forename: Shawn P
authors.initials: SP
authors.lastname: Williams
authors.forename: Andrea H
authors.initials: AH
authors.lastname: Ayscue
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Leesnitzer
authors.forename: G Bruce
authors.initials: GB
authors.lastname: Wisely
authors.forename: Andrew N
authors.initials: AN
authors.lastname: Billin
mesh_terms: D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D009154:Mutation; D057105:Nuclear Receptor Subfamily 4, Group A, Member 1; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D011987:Receptors, Steroid; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; D008024:Ligands; D057105:Nuclear Receptor Subfamily 4, Group A, Member 1; D018160:Receptors, Cytoplasmic and Nuclear; D011987:Receptors, Steroid; D014157:Transcription Factors