About this Event
25 South Green Drive, Athens, Ohio 45701
Chemistry & Biochemistry Colloquium | Justin Holub, April 17
The Chemistry & Biochemistry colloquium series presents Justin Holub discussing "Monitoring ligand-mediated helix 12 transitions within the human estrogen receptor α using bipartite tetracysteine display" on April 17 from 4:10 to 5:05 p.m. in Walter 135.
Holub is Associate Professor of Chemistry and Biochemistry at Ohio University. The host is Eric Masson.
ABSTRACT: Strategies that inform on protein structural dynamics in native environments greatly enhance our understanding of protein function and can facilitate the development of new therapies to treat disease. The human estrogen receptor a (ERa) is a ligand-mediated transcription factor that is modulated by the natural steroid hormone 17b-estradiol (E2). Structurally, the ERa ligand-binding domain (ERa-LBD) adopts a well-folded globular structure comprised of 11 a-helices and a short b-strand. The globular portion of the ERa-LBD is believed to be relatively static; however, the C-terminal helix 12 (H12) is dynamic and can adopt different configurations depending on what ligand occupies the LBD. In this study, we applied fluorescent labeling coupled with bipartite tetracysteine display to surveil ligand-mediated helix H12 transitions within the ERa-LBD. Wild-type ERa-LBDs were mutated to express a tetracysteine (C4) motif that binds to a biarsenical pro-fluorescent dye (FlAsH) upon transition of H12 to a folded state. Interestingly, binding of an agonist was accompanied by a reduction in FlAsH fluorescence, indicating that the C4 motif becomes occluded when H12 is tightly associated with the LBD. Our results also indicate that H12 remains flexible in unliganded receptors and receptors bound to pure antagonists, allowing for significant increases in FlAsH fluorescence. Such observations made it possible to determine association rates and equilibrium binding constants for FlAsH to the ERa-LBD in the presence and absence of estrogenic ligands. We anticipate that this genetically-encodable assay will be useful for studying how estrogenic compounds influence structural organization of the ERa-LBD and may be applied to studying helix transitions within other members of the nuclear receptor superfamily.
Ref: R. Pokhrel, T. Tang, J. M. Holub. Org Biomol Chem., 2020, 18, 6063-71
0 people are interested in this event