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研究生:曹美玲
研究生(外文):Mei-ling Tsao
論文名稱:C型肝炎病毒蛋白質-F蛋白質與Prefoldin-2之間的交互作用對於Prefoldin正常功能之影響
論文名稱(外文):Interaction of hepatitis C virus F protein with Prefoldin-2 perturbs normal function of Prefoldin
指導教授:徐善慧徐善慧引用關係葉昭廷葉昭廷引用關係
指導教授(外文):Shan-hui HsuChau-Ting Yeh
學位類別:碩士
校院名稱:國立中興大學
系所名稱:生命科學院碩士在職專班
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:2006
畢業學年度:94
語文別:中文
論文頁數:38
中文關鍵詞:C型肝炎病毒蛋白質—F蛋白質
外文關鍵詞:Hepatitis C virus F proteinprefoldin 2
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C型肝炎病毒 (Hepatitis C virus ;HCV) 是重要的人類病原體,全世界約有3﹪的人口受到感染,而此病毒是造成重大肝臟疾病,如肝癌、肝硬化的重要原因之一。C型肝炎病毒為正向單股RNA病毒,其RNA基因組約9.6 kb,可被轉譯成約3000個氨基酸的polyprotein,此polyprotein再經由病毒本身和宿主細胞的蛋白酶作用,產生至少10種病毒蛋白質—core,E1,E2,p7,NS2,NS3,NS4A,NS4B,NS5A和NS5B。在本篇報告中,吾人所研究的病毒蛋白質—F蛋白質,為一新發現的C型肝炎病毒蛋白質,與core蛋白質一樣經由相同的基因序列進行轉譯,但由於ribosomal frameshift 的原因,而產生有別於core蛋白質的基因產物。為瞭解此病毒蛋白質之生化特性,吾人利用酵母菌雙雜交系統 (yeast two-hybrid system) ,篩選出一個蛋白質—prefoldin 2,與F蛋白質之間有交互作用,之後再利用共同免疫沉澱法 (co-immunoprecipitation) 及雷射掃描共軛焦顯微鏡 (Laser scanning confocal microscope) 等技術,證明F蛋白質與prefoldin 2之間確實有蛋白質交互作用 ( protein-protein interaction )。此外,為了瞭解F蛋白質與prefoldin 2之間的交互作用是否會影響prefoldin之正常功能—幫助actin或α-tubulin形成正確聚合體的能力,經由雷射掃描共軛焦顯微鏡與nondenaturing gel electrophoresis的結果分析,發現當F蛋白質表達時,native
α-tubulin 濃度降低,而nonnative (or nascent) α-tubulin濃度不變。此結果表示F蛋白質與prefoldin 2之交互作用會干擾prefoldin之正常功能,進而影響α-tubulin 之摺疊過程。
Hepatitis C virus (HCV) is an important human pathogen that affects 170 million people worldwide. The HCV genome is approximately 9.6 kb in length and encodes a polyprotein, which is cleaved by viral and cellular proteases to produce at least 10 mature viral protein products. Hepatitis C virus F protein (HCV-F) is a newly discovered HCV gene product that is expressed by translational ribosomal frameshift within the capsid coding sequence. Its biological function and antigenic properties are unknown. Using yeast two-hybrid system, we identified a chaperone protein, prefoldin 2, interacting with F protein of hepatitis C virus. We confirmed the interaction in vivo using recombinant protein by co-immunoprecipitation and laser scanning confocal microscopy. Furthermore, we investigated whether the interaction of F protein with prefoldin 2 would affect the function of the prefoldin, that is, its ability to mediate correct polymerization of actin and α-tubulin. Using laser scanning confocal microscope and nondenaturing gel electrophoresis, we found the amount of native α-tubulin, but not nonnative (or nascent)
α-tubulin, decreased in the presence of F protein. The result revealed the interaction of HCV F protein with Prefoldin 2 affected the normal function of prefoldin, and thus, the folding of α-tubulin.
ABSTRACT
INTRODUCTION………………………………………………..1
MATERIALS AND METHODS…………………………………4
RESULTS……………………………………………………….15
DISCUSSION…………………………………………………..18
REFERENCES………………………………………………….21
FIGURES……………………………………………………….25
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