Structure of the PPARα and -γ Ligand Binding Domain in Complex with AZ 242; Ligand Selectivity and Agonist Activation in the PPAR Family

Abstract

The complex of PPARalpha and PPARgamma with the dual specificity agonist AZ 242 highlights the conserved interactions required for receptor activation. Together with the NMR data, this suggests a general model for ligand activation in the PPAR family. A comparison of the ligand binding sites reveals a molecular explanation for subtype selectivity and provides a basis for rational drug design.

Effect of heterodimer partner RXRα on PPARγ activation function-2 helix in solution

“…Intriguingly, residues Y473 and K474 in the AF-2 helix also exhibit moderate chemical shift changes. This is unexpected from the crystal structures because the conformations of the AF-2 helix H12 and the C-terminal segment D475-L476 in both homodimer [13] and heterodimer [10] configurations are very similar (Fig. 3E).…”

“…The 13 C α , 13 CO and 13 C β chemical shift deviations from the random coil values after correction for the deuterium isotope effect [4] and the secondary structure determined by the chemical shift index (CSI) program [9] are shown in Fig. 2.…”

“…3A and 3B. Using Δδ RMSD = 0.13 ppm as the cutoff, which corresponds to Δδ HN = 0.05 ppm, Δδ N = 0.4 ppm and Δδ CO = 0.3 ppm, or Δδ RMSD = 0.08 ppm when Δδ CO is excluded due to unavailable 13 CO resonance assignment in the heterodimer, we mapped residues with chemical shift perturbations above and below the cutoff in red (moderate) and blue (small), respectively, on the crystal structure [10], as shown in Fig. 3C The red residues are primarily located in H9 and H10, the H8-H9 and H9-H10 loops and the C-terminal residue Y477, all of which are components of the heterodimer interface.…”

“…DNA sequencing analysis confirmed the pET15b-PPARγ LBD sequence. The unlabeled RXRα LBD, PPARγ LBD and U-[ 2 H, 15 N, 13 C]-enriched PPARγ LBD were expressed in Escherichia coli BL21-DE3 bacteria, isolated by Talon affinity chromatography, digested by thrombin to remove the N-terminal His 6 tag, and finally purified by gel filtration chromatography as described [4].…”

“…Chemical shift-derived secondary structure of GW1929-bound PPARγ LBD monomer. The 13 C α , 13 CO and 13 C β chemical shift deviations between the measured and random coil values and the chemical shift index (CSI) consensus are plotted versus the primary sequence at the top. The three CSI values 1, 0 and −1 designate α-helix (H1-H12), random coil and β-sheet (S1-S4), respectively.…”