The effect of antagonists on the conformational exchange of the retinoid X receptor alpha ligand-binding domain

摘要

The effect of retinoid X receptor (RXR) antagonists on the conformational exchange of the RXR ligand-binding domain(LBD) remains poorly characterized. To address this question, we used nuclear magnetic resonance spectroscopy to comparethe chemical shift perturbations induced by RXR antagonists and agonists on the RXRa LBD when partnered with itself as ahomodimer and as the heterodimeric partner with the peroxisome proliferator-activated receptor y (PPARy) LBD. Chemical shiftmapping on the crystal structure showed that agonist binding abolished a line-broadening effect caused by a conformationalexchange on backbone amide signals for residues in helix H3 and other regions of either the homo- or hetero-dimer, whereasbinding of antagonists with similar binding affinities failed to do so. A lineshape analysis of a glucocorticoid receptor-interactingprotein 1 NR box 2 coactivator peptide showed that the antagonists enhanced peptide binding to the RXRa LBD homodimer,but to a lesser extent than that enhanced by the agonists. This was further supported by a lineshape analysis of the RXRC-terminal residue, threonine 462 (T462) in the homodimer but not in the heterodimer. Contrary to the agonists, the antagonistsfailed to abolish a line-broadening effect caused by a conformational exchange on the T462 signal corresponding to the RXRaLBD -antagonist- peptide ternary complex. These results suggest that the antagonists lack the ability of the agonists to shiftthe equilibrium of multiple RXRa LBD conformations in favor of a compact state, and that a PPARy LBD-agonist complex canprevent the antagonist from enhancing the RXRa LBD-coactivator binding interaction.