| Project/Area Number |
11839005
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Institution | Okazaki National Research Institutes (2000)
Tokyo Institute of Technology (1999) |
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Principal Investigator |
WATANABE Hajieme Okazaki National Research Institutes, Associate Professor, 統合バイオサイエンスセンター, 助教授 (80212322)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥4,200,000 (Direct Cost: ¥4,200,000)
Fiscal Year 2000: ¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1999: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Keywords | Endocrine Disruptors / Affinity Purification / Phthalates / Triazol / ターゲット / アフィニティー |
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| Research Abstract |
In order to clarify the molecular mechanisms of endocrine disruptors, I purified and identified molecular targets of endocrine disruptors by affinity purification. Generally, it has been thought that hormone receptor is a target molecule of en docrine disruptors, all phenomena caused by endocrine disruptors could not be explained only by hormone receptors. In order to examine the possibility that there are other target molecules of endocrine disruptors, we tried to directly purify the target molecules by affinity purification.
As target molecules, we used phthalate and amitrol. These chemicals were modified to fix on latex beads. The beads and spacer were developed in our lab oratory to reduce non-specific binding of proteins. As epoxyl groups are exist on the beads, chemicals having nucleophilic group such as amino group can be covalently fixed on the latex.
We purified endocrine disruptors binding proteins from HeLa cells. Nuclear extract and cytoplasmic extract were prepared from cultured cells and used. As the result, we could detect some binding proteins when both chemicals were used as ligands. After large-scale purification of the binding proteins, I determined amino acid sequences of those proteins and those cDNAs were cloned. No w I am expressing these genes and trying to analyze their function.
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