Selective regulation of gene expression by an orthogonal estrogen receptor–ligand pair created by polar-group exchange Original Research Article

Abstract Background: The nuclear and steroid hormone receptors function as ligand-dependent transcriptional regulators in eukaryotes. Hormone receptors have been engineered to selectively respond to synthetic ligands and used as remote regulators of gene expression for the study of gene function and as potential regulators of gene therapies. Results: In this work, a new ligand–receptor engineering strategy called ‘polar-group exchange’ is used to create a mutant form of the estrogen receptor, ER(Glu353→Ala), which lacks a carboxyl group critical for high-affinity binding of estradiol, but is able to transactivate in response to nanomolar concentrations of a carboxylate-functionalized estrogen analog, ES8. ES8 activates ER(Glu353→Ala) at concentrations that do not appreciably activate the ‘wild-type’ receptor ER(wt). Two similar carboxylate-functionalized ligands, ES6 and ES7, do not induce transactivation function. Similar selectivities are observed in ligand-binding assays in vitro, which follow the trends predicted by molecular modeling. Conclusions: Polar-group exchange is an effective strategy for rationally engineering ligand–receptor pairs. The ER(E353A)/ES8 ligand–receptor pair should constitute a unique and functionally orthogonal ligand-dependent transcri