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研究生:張家瑗
研究生(外文):Chia-Yuan Chang
論文名稱:人類小熱休克蛋白質HSPB3與actin交互作用的探討
論文名稱(外文):Characterization of the Interaction Between Human Small Heat Shock Protein HSPB3 and Actin
指導教授:張晃猷
指導教授(外文):Hwan-You Chang
學位類別:碩士
校院名稱:國立清華大學
系所名稱:生命科學系
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2002
畢業學年度:90
語文別:中文
論文頁數:71
中文關鍵詞:小熱休克蛋白質
外文關鍵詞:small heat shock proteinHSPB3
相關次數:
  • 被引用被引用:1
  • 點閱點閱:100
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  • 下載下載:0
  • 收藏至我的研究室書目清單書目收藏:0
本實驗室先前得到屬於小熱休克蛋白質家族的HSPB3,並且利用融合瘤技術製得H569F單株抗體,藉以偵測此蛋白質在大白鼠的腎臟、腦、骨骼肌等處皆有分布,尤以心臟的表現量為最高。由於HSPB3與其他小熱休克蛋白質比起來,屬於較新發現之成員,對於其功能及特性的了解仍少,因此我們利用酵母菌雙雜交系統,以HSPB3再次對人類心臟cDNA庫進行篩選。在經由反覆的酵母菌雙雜交確認後,我們篩選出在肌肉細胞中大量表現,同時扮演著極重要角色的actin能與HSPB3進行交互作用。為了近一步定義出此二蛋白質間彼此交互作用可能的結構部位,我們也分別對二者進行突變,以缺失不同部位的蛋白質片段分別與對方完整的蛋白質分子進行酵母菌雙雜交的測試,發現HSPB3主要是利用N端第24到48個胺基酸的部位與actin進行交互作用。actin則是利用較靠近C端的第302到337個胺基酸的部份來與HSPB3互相作用。其中參與actin單體間聚合的第324-327個胺基酸經由定點突變,並未發現對蛋白質交互作用造成影響。因此,actin與HSPB3進行交互作用的確切位置仍然需要進一步的分析。最後,藉由本論文所得到之資訊,我們能夠初步了解HSPB3在肌肉細胞中可能具有其獨特的功能。

Our laboratory previously identified a cDNA clone encoding the small heat shock protein HSPB3. A hybridoma that produces monoclonal antibody specific to HSPB3 was also generated. HSPB3 showed an extremely high expression level by western blot analysis in rat heart tissue, and could be detected in a less extent in skeletal muscle, brain and kidney. To further understand the physiological function of HSPB3, we screened the human heart cDNA library with the yeast two-hybrid system and showed that actin, which is expressed at a high level and plays an important role in muscle tissue, can interact with HSPB3. To identify the protein interaction domain, we made a series of truncated mutation of these two proteins and assayed their interaction by using the yeast two-hybrid system. The results indicated that the region containing amino acids 24-48 of HSPB3 is critical for interaction with actin. In actin, the C-terminal portion, especially amino acids 302-337, is necessary for its interaction with HSPB3. Nevertheless, site-specific mutagenesis of the amino acids 324-327, which involved in actin polymerization did not affect the interaction. Therefore, the exact region required for the HSPB3-actin interaction remains to be identified. In conclusion, our results have provided basic information required for a deeper insight into the properties and the functional role of the small heat shock protein HSPB3.

中文摘要------------------------------------------------------Ⅰ
英文摘要------------------------------------------------------Ⅱ
縮寫表--------------------------------------------------------Ⅲ
目錄----------------------------------------------------------Ⅴ
一、 前言-------------------------------------------------1
二、 材料與方法-------------------------------------------9
三、 實驗結果--------------------------------------------20
四、 討論------------------------------------------------27
五、 參考文獻--------------------------------------------35
圖表----------------------------------------------------------43附錄----------------------------------------------------------65

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羅明德,私立長庚大學碩士論文,類小熱休克蛋白質cDNA的定序及其在細胞中的表現,中華民國八十七年。
吳欣蓉,國立清華大學碩士論文,SHSPL及RLP-1單株抗體之製備與應用,中華民國八十八年。
蔡靜宜,國立清華大學碩士論文,人類小熱休克蛋白質HSPB3之特性與功能分析,中華民國八十九年。

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