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Structural requirements for high affinity ligand binding by estrogen receptors: a comparative analysis of truncated and full length estrogen receptors expressed in bacteria, yeast, and mammalian cells.

Publication ,  Journal Article
Wooge, CH; Nilsson, GM; Heierson, A; McDonnell, DP; Katzenellenbogen, BS
Published in: Mol Endocrinol
June 1992

In order to better understand the structural requirements for effective high affinity binding of estrogens and antiestrogens by the human estrogen receptor (ER), a comparative study was undertaken in which we examined: 1) native ER from the MCF-7 ER-positive human breast cancer cell line; 2) full length ER expressed in yeast; 3) the ER hormone binding domain (amino acid residues 302-595) expressed in yeast; 4) a bacterially expressed protein A fusion product encoding a truncated ER (amino acid residues 240-595); and 5) a synthetic peptide encompassing amino acids 510-551 of the ER. The binding parameters studied included affinity, kinetics, structural specificity for ligands, and stability. Full length ER expressed in yeast was very similar to the MCF-7 ER in its affinity [dissociation constant (Kd), 0.35 +/- 0.05 nM], dissociation rate (t1/2, 3-4 h at 25 C), and structural specificity for both reversible and covalently attaching affinity ligands. While the truncated ER expressed in yeast was similar to MCF-7 ER in its specificity of ligand binding, it showed a slightly reduced affinity for estradiol (Kd, 1.00 +/- 0.17 nM). The bacterially expressed ER also had a lower affinity for estradiol (Kd, 1.49 +/- 0.16 nM), which may be due in part to an increase in the dissociation rate (t1/2, 0.5 h at 25 C). The attachment of covalent affinity ligands and structural specificity for a variety of reversible ligands was comparable in the bacterially expressed ER to that observed for the receptors expressed in MCF-7 cells and yeast.(ABSTRACT TRUNCATED AT 250 WORDS)

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Published In

Mol Endocrinol

DOI

ISSN

0888-8809

Publication Date

June 1992

Volume

6

Issue

6

Start / End Page

861 / 869

Location

United States

Related Subject Headings

  • Tamoxifen
  • Saccharomyces cerevisiae
  • Recombinant Fusion Proteins
  • Receptors, Estrogen
  • Protein Conformation
  • Protein Binding
  • Peptide Fragments
  • Neoplasms, Hormone-Dependent
  • Neoplasm Proteins
  • Ligands
 

Citation

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Wooge, C. H., Nilsson, G. M., Heierson, A., McDonnell, D. P., & Katzenellenbogen, B. S. (1992). Structural requirements for high affinity ligand binding by estrogen receptors: a comparative analysis of truncated and full length estrogen receptors expressed in bacteria, yeast, and mammalian cells. Mol Endocrinol, 6(6), 861–869. https://doi.org/10.1210/mend.6.6.1495491
Wooge, C. H., G. M. Nilsson, A. Heierson, D. P. McDonnell, and B. S. Katzenellenbogen. “Structural requirements for high affinity ligand binding by estrogen receptors: a comparative analysis of truncated and full length estrogen receptors expressed in bacteria, yeast, and mammalian cells.Mol Endocrinol 6, no. 6 (June 1992): 861–69. https://doi.org/10.1210/mend.6.6.1495491.

Published In

Mol Endocrinol

DOI

ISSN

0888-8809

Publication Date

June 1992

Volume

6

Issue

6

Start / End Page

861 / 869

Location

United States

Related Subject Headings

  • Tamoxifen
  • Saccharomyces cerevisiae
  • Recombinant Fusion Proteins
  • Receptors, Estrogen
  • Protein Conformation
  • Protein Binding
  • Peptide Fragments
  • Neoplasms, Hormone-Dependent
  • Neoplasm Proteins
  • Ligands