碩士 / 國立海洋大學 / 水產生物技術研究所 / 87 / Abstract
Alzheimer's disease (AD) is the most common case of dementia occurring in mid- to late-life. Amyloid-b peptide (Ab) is the principal component of the extracellular deposits in AD. Ab promotes neurite outgrowth, generates reactive oxygen intermediates, induces cytotoxic cellular oxidant stress, and promotes microglial activation. However, Ab is derived from the transmembrane protein amyloid precursor protein (APP). A role for APP in the development of AD has been extensively researched in the genetic and cellular approach.
Reports obtained from different cell lines have shown that the secreted or membrane-associated forms of APP regulate cell growth, neurite out-growth and cell matrix adhesion. Nevertheless, function of APP and the relationship between APP and AD remain to be uncovered. Here we focus a acidic-rich domain (amino acid 188-220) in sAPP on the impact of biological activities in glial cell. For functional expression in E. coli, the acidic-rich domain was fused with thioredoxin (Trx) and cloned into the expression vector pET-23a. The expression vectors harboring the acidic-rich domain, rAPP; or Trx ; were transformed into BL21(DE3)pLysS. The transformants were induced with IPTG for overexpression, and the recombinant proteins were purified by ion-exchange chromatography.
The recombinant Trx-rAPP187-288 were demonstrated the biological activity that induced apoptosis of glial cells including RBA-1, C6 glioma and BV-2 microglial. The activating domain, between residue 220 to residue 288, was identified by deletion experiment of APP187-288. The recombinant APP220-288 showing EC50 = 1 μM were demonstrated a potent apoptotic activity. In this report we rule out the possibility that the apoptotic activity was resulted from the contaminant of LPS or creating a novel new protein by fusion with Trx. The apoptotic activity was enhanced by the pretreatment of the recombinant Trx-rAPP220-288, suggesting that apoptotic activity of rAPP may result in the interference of cell adhesion. Our results show a new finding for APP220-288 biological activity, inducing glial cell apoptosis, that may play an important role in Alzheimer's Disease.
| Identifer | oai:union.ndltd.org:TW/087NTOU0613008 |
| Date | January 1999 |
| Creators | Mei Jane Chuang, 莊美珍 |
| Contributors | Shye Jye Tang, 唐世杰 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 68 |
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