Structure, Energetics, and Dynamics of Binding Coactivator Peptide to the Human Retinoid X Receptor alpha Ligand Binding Domain Complex with 9-cis-Retinoic Acid

Abstract

Retinoid X receptors (RXRs) are ligand-dependent nuclear receptors, which are activated by the potent agonist 9-cis-retinoic acid (9cRA). 9cRA binds to the ligand binding domain (LBD) of RXRs and recruits coactivator proteins for gene transcription. Using isothermal titration calorimetry, the binding of a 13-mer coactivator peptide, GRIP-1, to the hRXR alpha-LBD homodimer complex containing 9cRA (hRXR alpha-LBD:9cRA:GRIP-1) is reported between 20 and 37 degrees C. Delta G is temperature independent (-8.5 kcal/mol), and GRIP-1 binding is driven by Delta H (-9.2 kcal/mol) at 25 degrees C. Delta C-p is large and negative (-401 cal mol(-1) K-1). The crystal structure of hRXR alpha-LBD:9cRA:GRIP-1 is reported at 2.05 angstrom. When the structures of hRXR alpha-LBD:9cRA:GRIP-1 and hRXR alpha-LBD:9cRA (1FBY) homodimers are compared, E453 and E456 on helix 12 bury and form ionic interactions with GRIP-1. R302 on helix 4 realigns to form new salt bridges to both E453 and E456. F277 (helix 3), F437 (helix 11), and F450 (helix 12) move toward the hydrophobic interior. The changes in the near-UV spectrum at 260 nm of the hRXR alpha-LBD:9cRA:GRIP-1 support this structural change. Helix 11 tilts toward helix 12 by approximate to 1 angstrom, modifying the ring conformation of 9cRA. Hydrogen-deuterium exchange mass spectroscopy indicates GRIP-1 binding to hRXR alpha-LBD:9cRA significantly decreases the exchange rates for peptides containing helices 3 (F277), 4 (R302), 11 (F437), and 12 (E453, E456). The structural changes and loss of dynamics of the GRIP-1-bound structure are used to interpret the energetics of coactivator peptide binding to the agonist-bound hRXR alpha-LBD.