Structural basis for PPAR partial or full activation revealed by a novel ligand binding mode

Abstract

The peroxisome proliferator-activated receptors (PPARs) are nuclear receptors involved in the regulation of the metabolic homeostasis and therefore represent valuable therapeutic targets for the treatment of metabolic diseases. The development of more balanced drugs interacting with PPARs, devoid of the side-effects showed by the currently marketed PPARγ full agonists, is considered the major challenge for the pharmaceutical companies. Here we present a structure-based virtual screening approach that let us identify a novel PPAR pan-agonist with a very attractive activity profile and its crystal structure in the complex with PPARα and PPARγ, respectively. In PPARα this ligand occupies a new pocket whose filling is allowed by the ligand-induced switching of the F273 side chain from a closed to an open conformation. The comparison between this pocket and the corresponding cavity in PPARγ provides a rationale for the different activation of the ligand towards PPARα and PPARγ, suggesting a novel basis for ligand design.

Figure 1. Flow chart of the SBVS strategy implemented in this work.

Figure 2. Dendrogram derived by clustering SIFts.

(A) Dendrogram derived by agglomerative hierarchical clustering of SIFt of PPARγ partial agonists and VS hits. Tanimoto similarity coefficient was used to calculate similarity between the SIFts. (B) Binding mode of VS hits belonging to cluster 1 (left), cluster 2 (middle) and cluster 3 (right). Some representatives of structures are shown.

Figure 3. Ligand binding to PPARα and PPARγ.

(A) Binding mode of AL29-26 (cyan) in the LBD of PPARγ (grey). (B) Binding mode of AL29-26 (yellow) in the LBD of PPARα (cyan); 2Fo-Fc omit maps are shown in mesh and contoured at 1σ. (C) Superposition of PPARα/AL29-26 (ligand yellow, protein cyan) and PPARγ/AL29-26 (ligand cyan, protein gray) crystal structures.

Figure 4. Comparison of PPARγ complexes.

(A) Superposition of PPARγ complexes (gray) with known partial agonists (pdb codes: 3D6D, 4PVU, 4PWL, 4JL4, 4JAZ, 4E4K, 2Q5P, 2Q6S, 2Q5S, 4E4Q, 5F9B) onto the PPARγ complex with AL29-26 (cyan). The carboxylate groups are depicted in red, the residue S342 in blue. (B) Superposition of SR2067 (magenta) (pdb code 4R06) onto the PPARγ complex with AL29-26 (cyan). The red arrow indicates the naphthalene groups of the two ligands.

Figure 5. Comparison of PPARα complexes.

(A) Superposition of PPARα complexes (gray) with known partial agonists (pdb codes: 2REW, 4BCR, 1K7L, 3SP6, 3FEI, 2GTK, 3G8I, 3ET1, 3KDT, 1I7G) onto the PPARα complex with AL29-26 (ligand yellow, protein cyan). The ligand BMS-631707 (PDB code 2REW) is shown in green. The “closed” (trans) conformation of F273 side-chain is also shown (gray). (B) New conformation of the loop 11–12 in the PPARα/AL29-26 complex: superposition of the loops 11–12 of known PPARα structures (light-brown) (same pdb codes of Fig. 5A) with that of PPARα/AL29-26 (ligand yellow, protein cyan). The ligand BMS-631707 (PDB code 2REW) is shown in green. Additional vdW interactions realized by AL29-26 are shown as red dashed lines.

Figure 6. New PPARα and PPARγ hydrophobic pockets.

(A) New PPARα (left) and PPARγ (right) hydrophobic pockets allowed by the ligand-induced switching of the F273 side-chain (F282 in PPARγ). (B) Modelled AL29-26 (gray) onto LT175 (green) (pdb code 3B3K) in the complex with PPARγ. Residues belonging to the “diphenyl pocket” are shown in red (vdW interactions with AL29-26 as red dashed lines). The residues interacting with the carboxylate group are shown in green (H-bonds are shown as blue dashed lines). (C) Superposition of PPARγ-modelled AL29-26 (gray) onto the PPARα/AL29-26 complex (the ligand is shown in yellow). Two representative residues of PPARγ are shown in gray, the corresponding residues of PPARα in cyan.