臺灣博碩士論文加值系統

C型肝炎病毒是一個正向單股病毒，是造成非A非B型肝炎的主要病因，並且可能發展成肝硬化甚至肝癌，然而有關於C型肝炎病毒致病機制並不清楚。已知C型肝炎病毒非結構蛋白質NS5B是病毒複製過程中重要的複製酶。雖然NS5B與病毒基因組的3’端具特異性結合，但它不具有模板選擇性，因此NS5B可能需要其他病毒蛋白質或宿主細胞蛋白質共同參與病毒複製。以酵母菌雙向雜交系統，利用HCV NS5B蛋白質尋找肝臟細胞中可能與其結合的蛋白質，篩選出的8個候選細胞因子當中，2個候選細胞因子以試管內共同免疫沉澱法，證明它們分別會與NS5B產生交互作用。
細胞因子PTB已證實可與C型肝炎病毒的3’端非轉譯區X區域和5’端非轉譯區的RNA結合，並調控C型肝炎病毒的複製與轉譯作用進行。本研究證明PTB與NS5B具交互作用，並進一步探討兩者間交互結合的區域。

The hepatitis C virus (HCV) is a positive single-stranded RNA virus and is an important pathogen for non-A, non-B hepatitis. HCV infection may sequentially induce chronic hepatitis to cirrhosis and hepatocellular carcinoma. Until now, the mechanisms of HCV pathogenesis have not yet completely understood. The HCV NS5B has been shown recently to serve as replicase in HCV replication. Although NS5B specifically binds to the 3’-end of the HCV genome, it does not display template specificity by itself. These results indicate that other viral or cellular protein(s) may involve in the replication of HCV. Using yeast two-hybrid system, eight potential NS5B interacting cellular clones were selected. Two of the selected clones were further shown the interacting property by in vitro co-immunoprecipitation assay.
Polypyrimidine binding protein (PTB) was shown to bind the 5’-untranslated region (UTR) and the 3’-end 98 nucleotide (X region) of HCV RNA. Both of these PTB-binding sites were thought to regulate HCV replication and translation. Here we show that HCV NS5B and PTB directly interact with each other in vitro. In addition, the interacting region(s) between NS5B and PTB were further determined.

目錄
中文摘要 i
英文摘要 ii
符號/縮寫 iii
致謝 v
第一章 導論 2
第二章 以酵母菌雙向雜交系統探討C型肝炎非結構性蛋白與細胞因
子的交互作用 8
2.1 概論 8
2.2 材料與方法 9
2.3 結果 16
2.4 討論 19
2.5 本章總結 21
第三章 探討C型肝炎非結構性蛋白與細胞因子PTB(Pyrimdine tract
binding protein)的交互作用 22
3.1 概論 22
3.2 材料與方法 23
3.3 結果 30
3.4 討論 35
3.5 本章總結 36
第四章 結論 37
附錄1. 圖表 38
附錄2. 質體建構流程圖 52
附錄3. 參考文獻 62

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