Date of Award

Spring 5-1999

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Chemistry

First Advisor

Dr. Wesley Gray

Second Advisor

Dr. Robert Miller

Third Advisor

Dr. Talmage Bursh

Abstract

The purpose of this study was to test the hypothesis that effects of xenoestrogens may be mediated through a low-affinity estrogen-binding site. This low-affinity estrogen-binding site (EBS) is found in rat uterus. A second receptor, the Type I-(high-affinity-EBS). is also present in the uterus and other tissues. In order to obtain the estrogen-binding fraction-from the uterus, an ammonium sulfate (AS) fractionation was performed on rat-uterine cytosol. Two fractions were made, a 40% AS, which contained the Type I estrogen receptor (ER) and a 70¾ AS fraction that contained the low-affinity estrogen-binding site. These fractions were assayed for their ability to bind to xenoestrogens. Binding assays performed on the 40% and 70% fractions show that 21 % of the activity is contained in the low-affinity estrogen-binding fraction, whereas only 7% of the activity was found in the Type I ER. These percentages are similar to those reported in the literature. Further analysis shows that xenoestrogens bind to the low-affinity estrogen-binding fraction with high affinity and specificity. The ligands, diethylstilbesterol (DES), bisphenol-A (BPA), and endosulfan (ENDO), were assayed for their ability to bind to the low-affinity estrogen-binding fraction. DES and BPA were shown to displace.,50% of the estradiol from the receptor. The ability of BPA to bind to the estrogen-binding fraction and Type I ER was examined. We found that BP A had a greater affinity for the low-affinity estrogen-binding site, as compared to the Type I. The low-affinity estrogen-binding fraction showed 20 fold higher affinity for BP A. These data suggest that the low-affinity estrogen-binding site may be the mediator of xenoestrogen action. Thus, our data indicates that the low-,affinity estrogen-binding site, which we believe to be a Type Il estrogen receptor, may play a greater role in cancer than previously predicted.

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