Thyroxine-thyroid hormone receptor interactions

ABSTRACTThyroid hormone (TH) actions are mediated by nuclear receptors (TRs α and β) that bind triiodothyronine (T3, 3,5,3′-triiodo-l-thyronine) with high affinity, and its precursor thyroxine (T4, 3,5,3′,5′-tetraiodo-l-thyronine) with lower affinity. T4 contains a bulky 5′ iodine group absent from T3. Because T3 is buried in the core of the ligand binding domain (LBD), we have predicted that TH analogues with 5′ substituents should fit poorly into the ligand binding pocket and perhaps behave as antagonists. We therefore examined how T4 affects TR activity and conformation. We obtained several lines of evidence (ligand dissociation kinetics, migration on hydrophobic interaction columns, and non-denaturing gels) that TR-T4 complexes adopt a conformation that differs from TR-T3 complexes in solution. Nonetheless, T4 behaves as an agonist in vitro (in effects on coregulator and DNA binding) and in cells, when conversion to T3 does not contribute to agonist activity. We determined x-ray crystal structures of the TRβ LBD in complex with T3 and T4 at 2.5-Å and 3.1-Å resolution. Comparison of the structures reveals that TRβ accommodates T4 through subtle alterations in the loop connecting helices 11 and 12 and amino acid side chains in the pocket, which, together, enlarge a niche that permits helix 12 to pack over the 5′ iodine and complete the coactivator binding surface. While T3 is the major active TH, our results suggest that T4 could activate nuclear TRs at appropriate concentrations. The ability of TR to adapt to the 5′ extension should be considered in TR ligand design

Low resolution structures of the retinoid X receptor DNA-binding and ligand-binding domains revealed by synchrotron x-ray solution scattering

Nuclear receptors are ligand-inducible transcription factors that share structurally related DNA-binding (DBD) and ligand-binding (LBD) domains. Biochemical and structural studies have revealed the modular nature of DBD and LBD. Nevertheless, the domains function in concert in vivo. While high-resolution crystal structures of nuclear receptor DBDs and LBDs are available, there are no x-ray structural studies of nuclear receptor proteins containing multiple domains. We report the solution structures of the human retinoid X receptor DBD-LBD (hRXR AB) region. We obtained ab initio shapes of hRXR AB dimer and tetramer to 3.3 and 1.7 nm resolutions, respectively, and established the position and orientation of the DBD and LBD by fitting atomic coordinates of hRXR DBD and LBD. The dimer is U-shaped with DBDs spaced at 2 nm in a head to head orientation forming an angle of about 10° with respect to each other and with an extensive interface area provided by the LBD. The tetramer is a more elongated X-shaped molecule formed by two dimers in head to head arrangement in which the DBDs are extended from the structure and spaced at about 6 nm. The close proximity of DBDs in dimers may facilitate homodimer formation on DNA; however, for the homodimer to bind to a DNA element containing two directly repeated halfsites, one of the DBDs would need to rotate with respect to the other element. By contrast, the separation of DBDs in the tetramers may account for their decreased ability to recognize DNA

Journal of Steroid Biochemistry and Molecular Biology

Trabalho completo: acesso restrito, p. 125–131Some nuclear receptor (NR) ligands promote dissociation of radiolabeled bound hormone from the buried ligand binding cavity (LBC) more rapidly than excess unlabeled hormone itself. This result was interpreted to mean that challenger ligands bind allosteric sites on the LBD to induce hormone dissociation, and recent findings indicate that ligands bind weakly to multiple sites on the LBD surface. Here, we show that a large fraction of thyroid hormone receptor (TR) ligands promote rapid dissociation (T1/2 < 2 h) of radiolabeled T3 vs. T3 (T1/2 ≈ 5–7 h). We cannot discern relationships between this effect and ligand size, activity or affinity for TRβ. One ligand, GC-24, binds the TR LBC and (weakly) to the TRβ-LBD surface that mediates dimer/heterodimer interaction, but we cannot link this interaction to rapid T3 dissociation. Instead, several lines of evidence suggest that the challenger ligand must interact with the buried LBC to promote rapid T3 release. Since previous molecular dynamics simulations suggest that TR ligands leave the LBC by several routes, we propose that a subset of challenger ligands binds and stabilizes a partially unfolded intermediate state of TR that arises during T3 release and that this effect enhances hormone dissociation

Differential effects of TR ligands on hormone dissociation rates: Evidence for multiple ligand entry/exit pathways

Some nuclear receptor (NR) ligands promote dissociation of radiolabeled bound hormone from the buried ligand binding cavity (LBC) more rapidly than excess unlabeled hormone itself This result was interpreted to mean that challenger ligands bind allosteric sites on the LBD to induce hormone dissociation, and recent findings indicate that ligands bind weakly to multiple sites on the LBD surface. Here we show, that a large fraction of thyroid hormone receptor (TR) ligands promote rapid dissociation (T(1/2) < 2 h) of , radiolabeled T(3) vs. T(3) (T(1/2), approximate to 5-7 h). We cannot discern relationships between this effect and ligand size, activity or affinity for TR beta. One ligand, GC-24, binds the TR LBC and (weakly) to the TR beta-LBD surface that mediates dimer/heterodimer interaction, but we cannot link this interaction to rapid T(3) dissociation. Instead, several lines of evidence suggest that the challenger ligand must interact with the buried LBC to promote rapid T(3) release. Since previous molecular dynamics simulations suggest that TR ligands leave the LBC by several routes, we propose that a subset of challenger ligands binds and stabilizes a partially unfolded intermediate state of TR that arises during T(3) release and that this effect enhances hormone dissociation. (C) 2009 Elsevier Ltd. All rights reserved.NIH[DK41482]U.S. National Institutes of Health (NIH)NIH[DK51281]U.S. National Institutes of Health (NIH)NIH[DK52798]U.S. National Institutes of Health (NIH)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)FAPESP[2006/00182-8