Monitoring Solution Structures of Peroxisome Proliferator-Activated Receptor ?/? upon Ligand Binding.

ABSTRACT: Peroxisome proliferator-activated receptors (PPARs) have been intensively studied as drug targets to treat type 2 diabetes, lipid disorders, and metabolic syndrome. This study is part of our ongoing efforts to map conformational changes in PPARs in solution by a combination of chemical cross-linking and mass spectrometry (MS). To our best knowledge, we performed the first studies addressing solution structures of full-length PPAR-?/?. We monitored the conformations of the ligand-binding domain (LBD) as well as full-length PPAR-?/? upon binding of two agonists. (Photo-) cross-linking relied on (i) a variety of externally introduced amine- and carboxyl-reactive linkers and (ii) the incorporation of the photo-reactive amino acid p-benzoylphenylalanine (Bpa) into PPAR-?/? by genetic engineering. The distances derived from cross-linking experiments allowed us to monitor conformational changes in PPAR-?/? upon ligand binding. The cross-linking/MS approach proved highly advantageous to study nuclear receptors, such as PPARs, and revealed the interplay between DBD (DNA-binding domain) and LDB in PPAR-?/?. Our results indicate the stabilization of a specific conformation through ligand binding in PPAR-?/? LBD as well as full-length PPAR-?/?. Moreover, our results suggest a close distance between the N- and C-terminal regions of full-length PPAR-?/? in the presence of GW1516. Chemical cross-linking/MS allowed us gaining detailed insights into conformational changes that are induced in PPARs when activating ligands are present. Thus, cross-linking/MS should be added to the arsenal of structural methods available for studying nuclear receptors.

SUBMITTER: Schwarz R 

PROVIDER: S-EPMC4798536 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature