Hydrogen/deuterium‐exchange (H/D‐Ex) of PPARγ LBD in the presence of various modulators
- 1 August 2006
- journal article
- Published by Wiley in Protein Science
- Vol. 15 (8) , 1883-1892
- https://doi.org/10.1110/ps.062103006
Abstract
A nuclear receptor, peroxisome proliferator-activated receptor gamma (PPARgamma), is a ligand-dependent transcription factor involved in glucose homeostasis and adipocyte differentiation. PPARgamma is the molecular target of various natural and synthetic molecules, including anti-diabetic agents such as rosiglitazone. Amide hydrogen/deuterium-exchange (H/D-Ex), coupled with proteolysis and mass spectrometry, was applied to study the dynamics of the PPARgamma ligand binding domain (LBD) with or without molecules that modulate PPARgamma activity. The H/D-Ex patterns of ligand-free PPARgamma LBD show that the ligand binding pocket of LBD is significantly more dynamic than the rest of the LBD. Presumably, the binding pocket is intrinsically disordered in order to accommodate different ligands. The presence of two full agonists (rosiglitazone and GW1929), a partial agonist (nTZDpa), and a covalent antagonist (GW9662), changed the dynamics/conformation of PPARgamma LBD and slowed the H/D exchange rate in various regions of the protein. The full agonists slowed the H/D exchange more globally and to a greater extent than the partial agonist or the antagonist, indicating that the full agonist stabilizes the PPARgamma LBD more than the partial agonist or the antagonist. One interesting observation is that the two full agonists significantly stabilized helix 12 while the partial agonist and the antagonist did not perturb the H/D exchange of this region. The results showed that the change in protein dynamics induced by ligand binding may be an important factor for the activation of genes and that H/D-Ex is a useful method for analyzing the biological activity of drug leads.Keywords
This publication has 35 references indexed in Scilit:
- Discovery and Characterization of a Substrate Selective p38α InhibitorBiochemistry, 2004
- Mechanism of the nuclear receptor molecular switchTrends in Biochemical Sciences, 2004
- Dynamics and Ligand-Induced Solvent Accessibility Changes in Human Retinoid X Receptor Homodimer Determined by Hydrogen Deuterium Exchange and Mass SpectrometryBiochemistry, 2004
- The Three-dimensional Structure of the Liver X Receptor β Reveals a Flexible Ligand-binding Pocket That Can Accommodate Fundamentally Different LigandsJournal of Biological Chemistry, 2003
- Amide Hydrogen Exchange Determined by Mass Spectrometry: Application to Rabbit Muscle AldolaseBiochemistry, 1996
- The nuclear receptor superfamily: The second decadeCell, 1995
- Primary structure effects on peptide group hydrogen exchangeProteins-Structure Function and Bioinformatics, 1993
- Determination of amide hydrogen exchange by mass spectrometry: A new tool for protein structure elucidationProtein Science, 1993
- Hydrogen exchange and structural dynamics of proteins and nucleic acidsQuarterly Reviews of Biophysics, 1983
- Protein dynamics investigated by the neutron diffraction–hydrogen exchange techniqueNature, 1982