Estrogen Receptor Binding of Xenoestrogens and Phytoestrogens in Japanese Quail (Coturnix japonica)
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Accession number;04A0112530
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| Title;Estrogen Receptor Binding of Xenoestrogens and Phytoestrogens in Japanese Quail (Coturnix japonica) |
| Author;
HANAFY A M
(Gifu Univ., Gifu, Jpn)
SASANAMI T
(Shizuoka Univ., Shizuoka, Jpn)
ICHIKAWA K
(Nagoya Univ., Nagoya, Jpn)
SHIMADA K
(Nagoya Univ., Nagoya, Jpn)
MORI M
(Shizuoka Univ., Shizuoka, Jpn)
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Journal Title;J Poult Sci
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Journal Code:G0008B
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ISSN:1346-7395
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VOL.41;NO.1;PAGE.30-37(2004)
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| Figure&Table&Reference;TBL.3, REF.22 |
| Pub. Country;Japan |
| Language;English |
| Abstract;Estrogenic compounds must bind to estrogen receptors (ER) and modulate estrogen-sensitive gene expression. However, various in vivo and in vitro assays have established that xenoestrogens and phytoestrogens are rather weak estrogens, with a 5,000- to 10,000-fold lower binding affinity to ER than estradiol-17.BETA. (E2). The purpose of this study was to determine the binding affinity of various estrogen-like chemicals to bacterially expressed quail ER.ALPHA.. The first-strand cDNA was synthesized from total RNA isolated from mature female quail liver with oligo-dT primed reverse transcription. The cDNA included the hinge region, the ligand-binding domain, and the C-terminal domain of quail ER.ALPHA.. It was amplified by PCR, and the PCR product was ligated into GST fusion protein expression vector, and transfected to the E. coli DH5.ALPHA. strain. A binding assay using the supernatant of the cell lysate was performed by incubation with [3H] E2 and increasing concentrations of competitor at 4.DEG.C. for 18h. Unbound steroids were removed by the addition of dextran-coated charcoal, followed by centrifugation for 15min. The radioactivity of the supernatant was determined with a liquid scintillation counter. Quail ER.ALPHA. expressed bacterially showed binding affinity to E2 with a dissociation constant of 1.74.+-.0.34X10-10M. The competition studies indicated that the relative binding affinities for the synthetic estrogens, diethylstilbestrol and ethynylestradiol, are very high, but that of the xenoestrogens, bisphenol A and nonylphenol, are very low. The phytoestrogens, coumestrol and genistein, can compete with E2 with a significant binding affinity. (author abst.) |
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