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研究生:沈雅倫
研究生(外文):Ya-Lun Shen
論文名稱:老鼠熱休克蛋白八十六之多聚體特性研究
論文名稱(外文):Oligomerization properties of Rat Heat Shock Protein 86
指導教授:張大慈
指導教授(外文):Margaret Dah-Tsyr Chang
學位類別:碩士
校院名稱:國立清華大學
系所名稱:分子與細胞生物研究所
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2004
畢業學年度:92
語文別:英文
論文頁數:74
中文關鍵詞:熱休克蛋白
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  • 被引用被引用:0
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熱休克蛋白九十 (HSP90) 是一種廣泛存於細胞質中的保護子,包括了兩種異構體α和β。HSP90α在老鼠中的同源基因(homologue)為HSP86,由733個胺基酸組成,可分為四個功能區:胺端區域 (第1到237個胺基酸),高電荷連接區域 (第238到272個胺基酸),中間區域 (第273到538個胺基酸) 和梭端區域 (第539到733個胺基酸)。HSP90的保護子功能是由ATP的水解和與其他輔保護子之交互作用調控。除此之外,升溫所引起的過渡狀態 – HSP90多聚體,是提高保護子功能的必要途徑。
我們利用三個HSP86的重組蛋白(HSP86-N, 第1到237個胺基酸;HSP86-NM, 第1到538個胺基酸;HSP86-C, 第539到733個胺基酸) 探究每個區域在形成多聚體時的特性。經由抑制檸檬酸合成酶沉澱測試,我們發現HSP86-N和HSP86-C 在成為多聚體之後,其保護子的功能上升了; 但在HSP86-NM卻反而降低。因此我們推測:在中間區域中,要形成多聚體的區位和檸檬酸合成脢結合的區位可能互相重疊,因而降低了它保護的功能。此外,就形成多聚體的臨界溫度 (Tm) 而言,胺端的臨界溫度小於梭端,顯示在全長的HSP86中,多聚體的形成應該是由胺端開始。梭端的多聚體會受ATP,ADP,鈣和鋅等因子影響,表示在梭端上有這些因子的結合區。
Heat shock protein 90 (HSP90) is an abundant chaperone in the cytosol, including two isoforms α and β. The rat homologue of HSP90 was HSP86, and defined as four domains in this study: N-terminal domain (a.a. 1~237), highly charged linker domain (a.a. 238~272), middle domain (a.a. 273~538) and C-terminal domain (a.a. 539~733). The chaperone activity of HSP90 is modulated by ATP hydrolysis and by interacting with several co-chaperones. In addition, heat-induced transition of HSP90 to self-oligomerization was also necessary for exerting the chaperone activity. Here, we used three HSP86 recombinants (HSP86N a.a. 1~237, HSP86NM a.a 1~538, HSP86C a.a 539~733) to explore the properties of oligomerization in each domain. From suppression assay, oligomerization of HSP86-N and HSP86-C resulted in higher chaperone function, whereas that of HSP86-NM showed lower chaperone function. We assumed that the oligomerization region in the middle domain overlapped with client binding domain. The N-terminal domain has lower critical temperature than the C-terminal domain, suggesting the first step of oligomerization of full length HSP86 starts from N-terminal. The oligomerization of HSP86-N and HSP86-C was affected by factors including ATP, ADP, Ca+ and Zn+, indicating there may be binding site for each factor.
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