Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγ
Peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear receptor transcription factor that regulates gene expression programs in response to ligand binding. Endogenous and synthetic ligands, including covalent antagonist inhibitors GW9662 and T0070907, are thought to compete for the or...
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eLife Sciences Publications Ltd
2024-11-01
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| Online Access: | https://elifesciences.org/articles/99782 |
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| author | Jinsai Shang Douglas J Kojetin |
| author_facet | Jinsai Shang Douglas J Kojetin |
| author_sort | Jinsai Shang |
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| description | Peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear receptor transcription factor that regulates gene expression programs in response to ligand binding. Endogenous and synthetic ligands, including covalent antagonist inhibitors GW9662 and T0070907, are thought to compete for the orthosteric pocket in the ligand-binding domain (LBD). However, we previously showed that synthetic PPARγ ligands can cooperatively cobind with and reposition a bound endogenous orthosteric ligand to an alternate site, synergistically regulating PPARγ structure and function (Shang et al., 2018). Here, we reveal the structural mechanism of cobinding between a synthetic covalent antagonist inhibitor with other synthetic ligands. Biochemical and NMR data show that covalent inhibitors weaken—but do not prevent—the binding of other ligands via an allosteric mechanism, rather than direct ligand clashing, by shifting the LBD ensemble toward a transcriptionally repressive conformation, which structurally clashes with orthosteric ligand binding. Crystal structures reveal different cobinding mechanisms including alternate site binding to unexpectedly adopting an orthosteric binding mode by altering the covalent inhibitor binding pose. Our findings highlight the significant flexibility of the PPARγ orthosteric pocket, its ability to accommodate multiple ligands, and demonstrate that GW9662 and T0070907 should not be used as chemical tools to inhibit ligand binding to PPARγ. |
| format | Article |
| id | doaj-art-fb8838a62f4b4e72936073ad314cfa87 |
| institution | Kabale University |
| issn | 2050-084X |
| language | English |
| publishDate | 2024-11-01 |
| publisher | eLife Sciences Publications Ltd |
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| series | eLife |
| spelling | doaj-art-fb8838a62f4b4e72936073ad314cfa872025-01-03T16:19:14ZengeLife Sciences Publications LtdeLife2050-084X2024-11-011310.7554/eLife.99782Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγJinsai Shang0https://orcid.org/0000-0001-8164-1544Douglas J Kojetin1https://orcid.org/0000-0001-8058-6168Department of Integrative Structural and Computational Biology, Scripps Research and The Herbert Wertheim UF Scripps Institute for Biomedical Innovation & Technology, Jupiter, United States; School of Basic Medical Sciences, Guangzhou Laboratory, Guangzhou Medical University, Guangzhou, ChinaDepartment of Integrative Structural and Computational Biology, Scripps Research and The Herbert Wertheim UF Scripps Institute for Biomedical Innovation & Technology, Jupiter, United States; Department of Biochemistry, Vanderbilt University, Nashville, United States; Center for Structural Biology, Vanderbilt University, Nashville, United States; Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, United States; Center for Applied AI in Protein Dynamics, Vanderbilt University, Nashville, United StatesPeroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear receptor transcription factor that regulates gene expression programs in response to ligand binding. Endogenous and synthetic ligands, including covalent antagonist inhibitors GW9662 and T0070907, are thought to compete for the orthosteric pocket in the ligand-binding domain (LBD). However, we previously showed that synthetic PPARγ ligands can cooperatively cobind with and reposition a bound endogenous orthosteric ligand to an alternate site, synergistically regulating PPARγ structure and function (Shang et al., 2018). Here, we reveal the structural mechanism of cobinding between a synthetic covalent antagonist inhibitor with other synthetic ligands. Biochemical and NMR data show that covalent inhibitors weaken—but do not prevent—the binding of other ligands via an allosteric mechanism, rather than direct ligand clashing, by shifting the LBD ensemble toward a transcriptionally repressive conformation, which structurally clashes with orthosteric ligand binding. Crystal structures reveal different cobinding mechanisms including alternate site binding to unexpectedly adopting an orthosteric binding mode by altering the covalent inhibitor binding pose. Our findings highlight the significant flexibility of the PPARγ orthosteric pocket, its ability to accommodate multiple ligands, and demonstrate that GW9662 and T0070907 should not be used as chemical tools to inhibit ligand binding to PPARγ.https://elifesciences.org/articles/99782NMR spectroscopynuclear receptorscovalent inhibitorpharmacologystructural biologycrystal structure |
| spellingShingle | Jinsai Shang Douglas J Kojetin Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγ eLife NMR spectroscopy nuclear receptors covalent inhibitor pharmacology structural biology crystal structure |
| title | Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγ |
| title_full | Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγ |
| title_fullStr | Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγ |
| title_full_unstemmed | Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγ |
| title_short | Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγ |
| title_sort | unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to pparγ |
| topic | NMR spectroscopy nuclear receptors covalent inhibitor pharmacology structural biology crystal structure |
| url | https://elifesciences.org/articles/99782 |
| work_keys_str_mv | AT jinsaishang unanticipatedmechanismsofcovalentinhibitorandsyntheticligandcobindingtopparg AT douglasjkojetin unanticipatedmechanismsofcovalentinhibitorandsyntheticligandcobindingtopparg |