Computational insights into the mechanism of ligand unbinding and selectivity of estrogen receptors.

Estrogen receptors (ER) belong to the nuclear receptor superfamily, and two subtypes, ERalpha and ERbeta, have been identified to date. The differentiated functions and receptor expressions of ERalpha and ERbeta made it attracted to discover subtype-specified ligands with high selectivity. However, these two subtypes are highly homologous and only two residues differ in the ligand binding pocket. Therefore, the mechanism of ligand selectivity has become an important issue in searching selective ligands of ER subtypes. In this study, steered molecular dynamics simulations were carried out to investigate the unbinding pathways of two selective ERbeta ligands from the binding pocket of both ERalpha and ERbeta, which demonstrated that the pathway between the H11 helix and the H7 approximately H8 loop was the most probable for ligand escaping. Then potentials of mean force for ligands unbinding along this pathway were calculated in order to gain insights into the molecular basis for energetics of ligand unbinding and find clues of ligand selectivity. The results indicated that His524/475 in ERalpha/ERbeta acted as a "gatekeeper" during the ligand unbinding. Especially, the H7 approximately H8 loop of ERbeta acted as a polar "transmitter" that controlled the ligand unbinding from the binding site and contributed to the ligand selectivity. Finally, the mechanism of ligand selectivity of ER subtypes was discussed from a kinetic perspective and suggestions for improving the ligand selectivity of ERbeta were also presented. These findings could be helpful for rational design of highly selective ERbeta ligands.