Aspartate 351 of estrogen receptor alpha is not crucial for the antagonistactivity of antiestrogens

dThe antagonist activity of antiestrogens is due to the presence of a long carbon side chain at positions 7 alpha or 11 beta or equivalent on their st eroid or steroid-like skeletons. These side chains establish hydrophobic in teractions with amino acids of the estrogen receptor alpha (ER alpha) ligan d binding domain. In addition, a hydrogen bond formed between amino acid As p-351 and the tertiary amine present at the end of the side chain of partia l antiestrogens is considered to be crucial for their antiestrogenicity. He re, we have investigated the role of Asp-351 in antiestrogen action in tran siently transfected HeLa and MDA-MB-231 cells. Our results indicate that di sruption of the negative charge at position 351 does not increase the agoni st activity of partial antiestrogens and thus that the hydrogen bond with t he antiestrogen side chain is not determinant in positioning the side chain in an antagonist position. The negative charge at position 351 was not req uired for transcriptional activity in the presence of hormone, but its pres ence was necessary for basal activity of the wild-type receptor and constit utive activities of mutants L536P and Y537A, suggesting a role of Asp-351 i n stabilizing the active conformation of ER alpha, 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 activ e conformation of the ER alpha ligand binding domain.