COOPERATIVITY AND DIMERIZATION OF RECOMBINANT HUMAN ESTROGEN-RECEPTORHORMONE-BINDING DOMAIN

The estrogen receptor dimerizes and exhibits cooperative ligand bindin g as part of its normal functioning. Interaction of the estrogen recep tor with its ligands is mediated by a C-terminal hormone-binding domai n (HBD), and residues within the HED are thought to contribute to dime rization, To examine dimer interactions in the isolated HBD, a human e strogen receptor HBD fragment was expressed in high yield as a cleavab le fusion protein in Escherichia coli, The isolated HBD peptide exhibi ted affinity for estradiol, ligand discrimination, and cooperative est radiol binding (Hill coefficient similar to 1.6) similar to the full-l ength protein. Circular dichroism spectroscopy suggests that the HBD c ontains significant amounts of alpha-helix (similar to 60%) and some b eta-strand (similar to 7%) and that ligand binding induces little chan ge in secondary structure. HBD dimer dissociation, measured using size exclusion chromatography, exhibited a half-life of similar to 1.2 h, which ligand binding increased similar to 3-fold (estradiol) to simila r to 4-fold (4-hydroxytamoxifen). These results suggest that the isola ted estrogen receptor HBD dimerizes and undergoes conformational chang es associated with cooperative ligand binding in a manner comparable t o the full-length protein, and that one effect of ligand binding is to alter the receptor dimer dissociation kinetics.