Conformational Diversity of the Helix 12 of the Ligand Binding Domain of PPARγ and Functional Implications.

Nuclear hormone receptors (NR) are transcription factors that activate gene expression in response to ligands. Structural and functional studies of the ligand binding domains (LBD) of NRs revealed that the dynamics of their C-terminal helix (H12) is fundamental for NR activity. H12 is rigid and facilitates binding of coactivator proteins in the agonist-bound LBD. In the absence of ligand, H12 exhibits increased flexibility. To provide a comprehensive picture of the H12 conformational equilibrium, extensive molecular dynamics simulations of the LBD of the PPARγ receptor in the presence or absence of ligand, and of coactivators and corepressor peptides, were performed. Free-energy profiles of the conformational variability of the H12 were obtained from more than four microseconds of simulations using adaptive biasing-force calculations. Our results demonstrate that, without ligand, multiple conformations of the H12 are accessible, including agonist-like conformations. We also confirm that extended H12 conformations are not accessible at ordinary temperatures. Ligand binding stabilizes the agonist H12 conformation relative to other structures, promoting a conformational selection. Similar effects are observed with coactivator association. The presence of corepressor peptides stabilizes conformations not allowed in the ligand-free, Rosiglitazone-bound or coactivator-bound LBDs. Corepressor binding, therefore, induces a conformational transition in the protein. Nevertheless, initial stages of corepressor dissociation could be induced by the ligand as it stabilizes the H12 in agonist form. Therefore, the present results provide a comprehensive picture of the H12 motions and their functional implications, with molecular resolution.