Monitoring ligand-mediated helix 12 transitions within the human estrogen receptor α using bipartite tetracysteine display

Estrogen receptor alpha ligand-binding domains (ER alpha-LBD) expressing tetracysteine motifs bind FlAsH-EDT(2)upon transition of helix 12 (H12) to a folded state. Changes in fluorescence intensity allowed surveillance of ligand-mediated H12 transitions and facilitated the determination of FlAsH association rates (k(on)) and apparent equilibrium dissociation constants (K-app) to ER alpha-LBDs in the presence of estrogenic ligands.