Aspartate 351 of Estrogen Receptor α Is Not Crucial for the Antagonist Activity of Antiestrogens

• Published in: The Journal of biological chemistry 2000-07, Vol.275 (27), p.20867-20872
• Main Authors: Anghel, Silvia I., Perly, Véronique, Melançon, Geneviève, Barsalou, Annie, Chagnon, Samuel, Rosenauer, Angelika, Miller, Wilson H., Mader, Sylvie
• Format: Article
• Language: English
• Subjects: Animals; Aspartic Acid - genetics; Aspartic Acid - metabolism; Binding Sites; COS Cells; Estradiol - pharmacology; Estrogen Receptor alpha; Estrogen Receptor Modulators - pharmacology; Gene Expression Regulation - drug effects; HeLa Cells; Humans; Mutation; Nuclear Receptor Coactivator 2; Protein Binding; Protein Conformation; Receptors, Estrogen - antagonists & inhibitors; Receptors, Estrogen - genetics; Receptors, Estrogen - metabolism; Transcription Factors - metabolism; Transfection
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M002098200

The antagonist activity of antiestrogens is due to the presence of a long carbon side chain at positions 7α or 11β or equivalent on their steroid or steroid-like skeletons. These side chains establish hydrophobic interactions with amino acids of the estrogen receptor α (ERα) ligand binding domain. In addition, a hydrogen bond formed between amino acid Asp-351 and the tertiary amine present at the end of the side chain of partial antiestrogens is considered to be crucial for their antiestrogenicity. Here, we have investigated the role of Asp-351 in antiestrogen action in transiently transfected HeLa and MDA-MB-231 cells. Our results indicate that disruption of the negative charge at position 351 does not increase the agonist activity of partial antiestrogens and thus that the hydrogen bond with the antiestrogen side chain is not determinant in positioning the side chain in an antagonist position. The negative charge at position 351 was not required for transcriptional activity in the presence of hormone, but its presence was necessary for basal activity of the wild-type receptor and constitutive activities of mutants L536P and Y537A, suggesting a role of Asp-351 in stabilizing the active conformation of ERα. This stabilizing role of Asp-351 could be due to interaction of Asp-351 with the amide group of the peptide bond between Leu-539 and Leu-540 in helix 12 observed in the active conformation of the ERα ligand binding domain.