臺灣博碩士論文加值系統

本研究將酵母菌中唯一的第一型精胺酸甲基轉移酶－Hmt1p，表現於大腸桿菌中。藉由膠體色層分析法得知，重組Hmt1p的構形取決於其濃度的高低，即濃度增加時，雙聚體的蛋白質便聚合成四或六聚體的構形。而體外實驗也指出，蛋白質N端的His6-tag及結構變化僅對其受質專一性造成些微的影響。經由兩段一級結構極相似的人工合成胜肰進行體外分析，証實Hmt1p是以nonprocessive的方式對受質進行甲基化作用。
將Hmt1p固定於affi-gel 10上，並與野生型酵母菌液行下拉作用以拖曳出其互動蛋白。利用胜肰質量指紋譜鑑定出22個蛋白質，包括：Ssb2p、Ssa1p/Ssa2p、Nic96p、Cdc19p、Ded1p、Pdc1p、Dbp2p、Tef1p、Hem15p、Tdh3p、Ydj1p、Rps3p、Rps4ap、Yra1p、Fmp52p、Rps7ap、Rps1bp、Rps8bp、Rps19ap/Rps19bp及Rps14bp。並利用[3H]AdoMet和固定式Hmt1p及其互動蛋白作用，以蛋白質電泳分離標誌出的受質並曝光至Ｘ光片辨視之，發現有兩條曝光帶出現於膠體上Rps4ap及Rps7ap極相近的位置上。同樣利用蛋白質電泳分離40S核糖體，並取得Rps4ap及Rps7ap，然後以質譜儀進行胺基酸分析。發現這兩個蛋白質皆不被甲基化，証實所觀察到的曝光帶應是來自其他的蛋白質，因此，尚需更多的實驗去辨視其身份。

目錄

目錄---------------------------------------------------- Ⅰ
英文摘要------------------------------------------------ Ⅱ
中文摘要------------------------------------------------ Ⅲ
表次---------------------------------------------------- Ⅳ
圖次---------------------------------------------------- Ⅴ
附錄文次------------------------------------------------ Ⅶ


緒論----------------------------------------------------- 1
實驗材料與方法------------------------------------------- 8
結果---------------------------------------------------- 28
討論---------------------------------------------------- 41
參考文獻------------------------------------------------ 52
附表---------------------------------------------------- 57
附圖---------------------------------------------------- 61
附錄---------------------------------------------------- 91

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