Ligand-mediated modulation of estrogen receptor conformation by estradiol analogs

JA Schwartz, DF Skafar - Biochemistry, 1993 - ACS Publications
JA Schwartz, DF Skafar
Biochemistry, 1993ACS Publications
Revised Manuscript Received July 13, 1993* abstract: The studies presented here show
how changing the structure of the ligand can affect the conformation of the receptor. Five
different estradiol analogs have been tested for binding tothe calf uterine estrogen receptor.
In three of the analogs the phenolic hydroxyl group had been moved from the 3 to the 1, 2, or
4 position on the A-ring (1-hydroxyestratrien-17| 8-ol, 2-hydroxyestratrien-17j8-ol, or 4-
hydroxyestratrien-173-ol). In the remaining two analogs either the A-or the D-ring hydroxyl …
Revised Manuscript Received July 13, 1993* abstract: The studies presented here show how changing the structure of the ligand can affect the conformation of the receptor. Five different estradiol analogs have been tested for binding tothe calf uterine estrogen receptor. In three of the analogs the phenolic hydroxyl group had been moved from the 3 to the 1, 2, or 4 position on the A-ring (1-hydroxyestratrien-17| 8-ol, 2-hydroxyestratrien-17j8-ol, or 4-hydroxyestratrien-173-ol). In the remaining two analogs either the A-or the D-ring hydroxyl group had been removed altogether (estratrien-17/9-ol or 3-hydroxyestratriene). Competition binding assay showed that the relative binding affinity for the estrogen receptor had been weakenedby all changes in the structure of the ligand. Furthermore, the ligands in which either the 3/3-or the 173-hydroxyl group was missing produced nonparallel slopes in the linear portions of the displacement curves compared to that of estradiol; the ligands in which the phenolic hydroxyl had simply been movedaround the A-ring, however, did not. These observations implied that the receptor binding mechanism used by the monohydroxyl ligands was different from that of estradiol. Saturation binding analysis showed that while the presence of any of the dihydroxyl ligands or that of estratrien-173-ol decreased the positive cooperativity of the [3H] estradiol-estrogen receptor interaction, the presence of the 3-hydroxyestratriene ligand increased it. These results suggest that both the binding mechanismand the affinity of the ligand for the receptor are exquisitely sensitive to the structure of the ligand.
The binding of estradiol to its receptor ultimately leads to the many observed biological responses to the hormone. One early step in this process is the production of conformational changes in the receptor by ligand binding. Previous studies have shown that two conformational states exist for the estrogen receptor which differ in their affinity for estradiol (Weichman & Notides, 1977). The formationof the high-affinity state is dependent on receptor dimerization and is characterized by positive cooperative estradiol binding, as demonstrated by a convex Scatchard plot and a Hill coefficient of 1.6 (Notides et al., 1981). Positivecooperativity is a type of self-regulation in which the binding of the first molecule of ligand to an oligomeric protein increases the affinity of the protein for additional molecules of ligand. The positive cooperative binding mechanism is then an indicator of conformational change in the protein. However, for the estrogen receptor, therelation between the structure of the ligand and itsbinding mechanism, that is, the presence and degree of positive cooperativity, has not been defined. Structure-activity studies show that many different com-pounds bind to the estrogen receptor; however, binding affinity is not necessarily related to estrogenicity (Jordan & Tate, 1980). Numerousefforts have been aimed atcorrelating the position of the twohydroxyl groups on the estradiol molecule to the binding affinity and the conformational changes elicited. Early works proposed that binding affinity was dependent on the A-ring hydroxyl group (Duax et al., 1980; Jordan, 1984), while conformational changes, correct folding, and stabilization of the dimeric receptor form were dependent on the D-ring hydroxyl (Vedeckis et al., 1980; Duax et al., 1980;
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