A structural mechanism for directing corepressor-selective inverse agonism of PPAR\u03b3

Theodore M Kamenecka,Andrew Cano,Anne-Laure Blayo,Douglas J Kojetin,Jinsai Shang,Ian M Chrisman,Zahra Heidari,Jared Bass,Michelle D Nemetchek,Richard Brust,Jakob Fuhrmann,Sarah A Mosure,Patrick R Griffin,Travis S Hughes

doi:10.1038/s41467-018-07133-w

Theodore M Kamenecka, Andrew Cano + Show 12 more

Open Access

https://doi.org/10.1038/s41467-018-07133-w

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Abstract

Small chemical modifications can have significant effects on ligand efficacy and receptor activity, but the underlying structural mechanisms can be difficult to predict from static crystal structures alone. Here we show how a simple phenyl-to-pyridyl substitution between two common covalent orthosteric ligands targeting peroxisome proliferator-activated receptor (PPAR) gamma converts a transcriptionally neutral antagonist (GW9662) into a repressive inverse agonist (T0070907) relative to basal cellular activity. X-ray crystallography, molecular dynamics simulations, and mutagenesis coupled to activity assays reveal a water-mediated hydrogen bond network linking the T0070907 pyridyl group to Arg288 that is essential for corepressor-selective inverse agonism. NMR spectroscopy reveals that PPARγ exchanges between two long-lived conformations when bound to T0070907 but not GW9662, including a conformation that prepopulates a corepressor-bound state, priming PPARγ for high affinity corepressor binding. Our findings demonstrate that ligand engagement of Arg288 may provide routes for developing corepressor-selective repressive PPARγ ligands.

Highlights

Small chemical modifications can have significant effects on ligand efficacy and receptor activity, but the underlying structural mechanisms can be difficult to predict from static crystal structures alone
In addition to their utility as nondissociative orthosteric competitive ligands that inhibit binding of other orthosteric PPARγ ligands, our data suggest that T00709707 and GW9662 can be classified as a covalent corepressor-selective inverse agonist and a covalent antagonist, respectively
One of these conformations is shared with GW9662-bound PPARγ, significantly populated by the R288A mutant that cannot form the pyridylwater-R288 network and shows similar backbone amide chemical shift values indicating a similar conformation to the coactivatorbound state

Summary

Introduction

Small chemical modifications can have significant effects on ligand efficacy and receptor activity, but the underlying structural mechanisms can be difficult to predict from static crystal structures alone. Originally it was thought that full activation of PPARγ was required for antidiabetic efficacy, recent studies have shown that antidiabetic PPARγ ligands alter posttranslational modifications impacting target gene expression These ligands have been shown to span a wide range of efficacies—including full and partial agonists, antagonists, and inverse agonists that have robust or mild activating, neutral, or repressive transcriptional properties, respectively[2,3,4,5]. Partial agonists generally do not hydrogen bond to Y473, but mildly stabilize helix 12 via interactions with other regions of the ligand-binding pocket, resulting in less pronounced changes in coregulator affinity and transcriptional activation[15,16,17]. NMR analysis shows that T0070907-bound PPARγ populates two long-lived structural conformations, one of which resembles the state populated by GW9662 and a unique state that is similar to the corepressor-bound state, revealing a structural mechanism for directing corepressor-selective repression of PPARγ

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