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Mechanistic elucidation guided by covalent inhibitors for the development of anti-diabetic PPAR? ligands.

ABSTRACT: Peroxisome proliferator-activated receptor gamma (PPAR?) is a ligand-regulated transcription factor that plays crucial roles in adipogenesis, lipid metabolism, and glucose homeostasis. Several PPAR? ligands possess anti-diabetic activity and they commonly inhibit the phosphorylation of PPAR? at serine 273 (Ser273). The recently reported PPAR? ligand SR1664, which selectively blocks the phosphorylation of PPAR? without classical agonism, has potent anti-diabetic activity, indicating that the inhibition of Ser273 phosphorylation is sufficient to provoke anti-diabetic effects. In this study, we revealed the X-ray structure of PPAR? co-crystallized with SR1664 bound to the alternate binding site of PPAR? and confirmed that the alternate site binding of SR1664 blocks the phosphorylation of Ser273. Furthermore, using covalent inhibitors as chemical tools, we demonstrated that the inhibition of phosphorylation is attributed to the occupation of a specific site which is a hydrophobic region between helix 3 and ?3-?4 at the binding pocket of PPAR?. In high-fat diet-induced obese mice, we confirmed the anti-diabetic activity of our covalent inhibitor SB1453 that was designed to bind at the specific site in PPAR? for blocking the phosphorylation of Ser273. Lastly, the target selectivity of SB1453 was demonstrated by fluorescence-based visualization of target proteins complexed with the covalent probe 11 containing a bioorthogonal functional group.

SUBMITTER: Bae H

PROVIDER: S-EPMC6021786 | biostudies-literature | 2016 Aug

REPOSITORIES: biostudies-literature

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Mechanistic elucidation guided by covalent inhibitors for the development of anti-diabetic PPARγ ligands.

Bae Hwan H Jang Jun Young JY Choi Sun-Sil SS Lee Jae-Jin JJ Kim Heejun H Jo Ala A Lee Kong-Joo KJ Choi Jang Hyun JH Suh Se Won SW Park Seung Bum SB

Chemical science 20160513 8

Peroxisome proliferator-activated receptor gamma (PPARγ) is a ligand-regulated transcription factor that plays crucial roles in adipogenesis, lipid metabolism, and glucose homeostasis. Several PPARγ ligands possess anti-diabetic activity and they commonly inhibit the phosphorylation of PPARγ at serine 273 (Ser273). The recently reported PPARγ ligand SR1664, which selectively blocks the phosphorylation of PPARγ without classical agonism, has potent anti-diabetic activity, indicating that the inhi ...[more]

PMID: 30034693

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Project description:The ubiquitin-proteasome system is responsible for the degradation of proteins and plays a critical role in key cellular processes. While the constitutive proteasome (cPS) is expressed in all eukaryotic cells, the immunoproteasome (iPS) is primarily induced during disease processes, and its inhibition is beneficial in the treatment of cancer, autoimmune disorders and neurodegenerative diseases. Oxathiazolones were reported to selectively inhibit iPS over cPS, and the inhibitory activity of several oxathiazolones against iPS was experimentally determined. However, the detailed mechanism of the chemical reaction leading to irreversible iPS inhibition and the key selectivity drivers are unknown, and separate characterization of the noncovalent and covalent inhibition steps is not available for several compounds. Here, we investigate the chemical reaction between oxathiazolones and the Thr1 residue of iPS by quantum mechanics/molecular mechanics (QM/MM) simulations to establish a plausible reaction mechanism and to determine the rate-determining step of covalent complex formation. The modelled binding mode and reaction mechanism are in line with the selective inhibition of iPS versus cPS by oxathiazolones. The kinact value of several ligands was estimated by constructing the potential of mean force of the rate-determining step by QM/MM simulations coupled with umbrella sampling. The equilibrium constant Ki of the noncovalent complex formation was evaluated by classical force field-based thermodynamic integration. The calculated Ki and kinact values made it possible to analyse the contribution of the noncovalent and covalent steps to the overall inhibitory activity. Compounds with similar intrinsic reactivities exhibit varying selectivities for iPS versus cPS owing to subtle differences in the binding modes that slightly affect Ki, the noncovalent affinity, and importantly alter kinact, the covalent reactivity of the bound compounds. A detailed understanding of the inhibitory mechanism of oxathiazolones is useful in designing iPS selective inhibitors with improved drug-like properties.

Dataset Information

Mechanistic elucidation guided by covalent inhibitors for the development of anti-diabetic PPAR? ligands.

Publications

Mechanistic elucidation guided by covalent inhibitors for the development of anti-diabetic PPARγ ligands.

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