Small-angle X-ray scattering studies on structures of an estrogen-related receptor alpha ligand binding domain and its complexes with ligands and coactivators.
We have investigated the structure of the mouse estrogen-related receptor alpha ligand binding domain (mERRalpha LBD) with a flexible hinge region (including more than 30 residues) in solution and the variations of its structure in response to binding with various ligands and coactivators by using synchrotron small-angle X-ray scattering (SAXS) and biochemical analysis. The mERRalpha LBD alone and in complex with the domain containing all three leucine-rich repeats of human peroxisome proliferator-activated receptor gamma coactivator-1alpha (hPGC-1alpha) were both cloned and expressed in Escherichia coli and purified to homogeneity. We also synthesized steroid receptor coactivator-1 (SRC-1), a 15-mer peptide corresponding to the leucine-rich repeat 4 of human SRC-1. We tested 4-hydroxytamoxifen, diethylstilbestrol, chlordane, genistein, daidzein, and biochanin A as ligands. SAXS and biochemical analyses were used to show that the apo mERRalpha LBD is present as a homodimer in solution. The apo mERRalpha LBD homodimer has an asymmetrically ellipsoidal shape with a lower region that includes the extended hinge domain. It was found that the homodimer always undergoes some structural changes in cooperation with the hinge domain when it binds with ligands. These structural changes are probably due to changes in the conformation of the mERRalpha LBD ligand binding pocket, which has a relatively small volume when ligand bindings occur. Overall, all the tested ligands have similar impacts on the structure of the mERRalpha LBD. In contrast, the mERRalpha LBD:hPGC-1alpha complex has a Y-shaped structure, which is quite different from the structures of the mERRalpha LBD bound with ligands. This result provides important information about the spatial orientation of the flexible disordered and/or relatively rigid regions bearing all three L1, L2, and L3 motifs of the partially unfolded hPGC-1alpha fragments, which are involved in the interaction with the mERRalpha LBD. The mERRalpha LBD was found to have a strong binding affinity with the hPGC-1alpha coactivator fragment, which arises from the cooperativity of the three leucine-rich motifs in the coactivator binding mode. In contrast, the mERRalpha LBD complex with SRC-1 was found to have a structure very similar to that of the apo mERRalpha LBD, which provides a foundation for a ligand-independent understanding of the constitutively transcriptional activity of apo ERRs.
Jin
Kyeong Sik
KS
National Research Laboratory for Polymer Synthesis and Physics, Pohang Accelerator Laboratory, Center for Integrated Molecular Systems, Polymer Research Institute, Pohang University of Science and Technology, Pohang, Republic of Korea.
Park
Joon Kyu
JK
Yoon
Jinhwan
J
Rho
Yecheol
Y
Kim
Jae-Hong
JH
Kim
Eunice EunKyeong
EE
Ree
Moonhor
M
D000595:Amino Acid Sequence; D001665:Binding Sites; D002942:Circular Dichroism; D047628:Estrogen Receptor alpha / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D053838:Scattering, Small Angle; D017386:Sequence Homology, Amino Acid; D014961:X-Ray Diffraction
D047628:Estrogen Receptor alpha; D008024:Ligands