The first structure of a muntidomain human estrogen receptor alpha (hERα).

Human estrogen receptor α plays a critical role in cell growth, and cancer metastasis. It belongs to a class of proteins called nuclear receptors (NR), which typically consist of a DNA binding domain (DBD) and a ligand binding domain (LBD). The physical interactions between the two domains are characteristic of the different NR proteins and have been found to be functionally significant. The structure of this inter- domain interface is therefore important to understand how hERα is induced by the ligand Estradiol to bind specific genes in order to regulate their expression. In order to acquire structural insights into the interaction between the DBD and LBD of hERα Dr. Sichun Yang’s team (Case Western Reserve University Department of Nutrition, School of Medicine) collaborated with BioCAT to acquire small angle x-ray scattering (SAXS) data, which was then combined with complementary techniques such as hydroxyl-radical foot-printing to get a picture of the key regions that have hydrophobic amino acid residues that mediate the interaction between the two domains, and the relative spatial orientation of the two domains in three dimensional space. The L-shaped structure that hERα showed in this study turned out to be a novel DBD-LBD conformation.

The size-exclusion chromatography SAXS (SEC-SAXS) instrument at BioCAT, which ensures the highest possible quality of the sample passing through the SAXS cell through a size based separation of the sample from aggregates and other contaminants has made on-site biochemical experiments extremely convenient and was put to optimal use in studying the architectural variations in hERα with and without the ligand. This work also demonstrates the power of combining different complementary techniques to create a more thorough and validated mechanistic and structural understanding of a biological macromolecular system. Modeling of data from hydroxyl-radical foot-printing and SAXS was aided by molecular dynamics approaches in a multi-technique platform called iSPOT (integration of Scattering, footPrinting, and dOcking simulaTion) developed by the Yang lab . Furthermore, a functional assay that involves measuring the fluorescence anisotropy of a dye-labeled DNA in response to varying concentrations of Estradiol bound hERα with selected mutations in the inter-domain interface shed valuable light on the mechanism by which changes in the LBD can ‘allosterically’ affect the structure and function of the DBD. Some of these mutations are naturally found in certain cancers and may be key to understanding metastasis and in the development of targeted therapeutic strategies.

See: Wei Huang, Yi Peng, Janna Kiselar, Xuan Zhao, Aljawharah Albaqami, Daniel Mendez, Yinghua Chen, Srinivas Chakravarthy, Sayan Gupta, Corie Ralston, Hung-Ying Kao, Mark R. Chance, Sichun Yang, “Multidomain architecture of estrogen receptor reveals interfacial cross-talk between its DNA-binding and ligand-binding domains,” Nat. Commun. 9 (3520), 3520-1-3520-10 (2018). DOI: 10.1038/s41467-018-06034-2