The peroxisome proliferator-activated receptor-[Formula: see text] (PPAR[Formula: see text]) is a member of PPAR nuclear receptor family, and its antagonists have been widely used to treat pediatric metabolic disorders. Traditional type-1 and type-2 PPAR[Formula: see text] antagonists are all small-molecule compounds that have been developed to target the ligand-binding site (LBS) of PPAR[Formula: see text], which is not overlapped with the coactivator-interacting site (CIS) of PPAR[Formula: see text]. In this study, we described the rational design of type-3 peptidic antagonists that can directly disrupt PPAR[Formula: see text]-coactivator interaction by physically competing with coactivator proteins for the CIS site. In the procedure, seven reported PPAR[Formula: see text] coactivator proteins were collected and eight 11-mer helical peptide segments that contain the core PPAR[Formula: see text]-binding LXXLL motif were identified in these coactivators, which, however, possessed a large flexibility and intrinsic disorder when splitting from coactivator protein context, and thus would incur a considerable entropy penalty (i.e. indirect readout) upon binding to PPAR[Formula: see text] CIS site. By carefully examining the natively folded conformation of these helical peptides in their parent protein context and in their interaction mode with the CIS site, we rationally designed a hydrocarbon bridge across the solvent-exposed, ([Formula: see text], [Formula: see text]+ 4) residues to constrain their helical conformation, thus largely minimizing the unfavorable indirect readout effect but having only a moderate influence on favorable enthalpy contribution (i.e. direct readout) upon PPAR[Formula: see text]-peptide binding. The computational findings were further substantiated by fluorescence competition assays.
Keywords: Peroxisome proliferator-activated receptor-α; biomolecular modeling; hydrocarbon stapling; pediatric metabolic disorder; type-3 peptidic antagonist.