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    Title: 吡啶釕金屬錯合物與大腸桿菌作用之蛋白質在二維電泳的分佈
    Other Titles: Protein distribution in two-dimensional polyacrylamide gel electrophoresis induced by polypyridyl ruthenium complexes
    Authors: 張芳英;Chang, Fang-ying
    Contributors: 淡江大學生命科學研究所碩士班
    鄭建中;Cheng, Chien-chung
    Keywords: 吡啶釕金屬錯合物;二維凝膠電泳;polypyridyl ruthenium complex;2-D PAGE
    Date: 2007
    Issue Date: 2010-01-11 02:29:13 (UTC+8)
    Abstract: 抗癌藥物釕金屬錯合物在臨床醫學上對治療腫瘤上具有相當的發展潛力。基於其毒性低、水溶性高及有高度特異選擇性等特點,可比美抗癌藥物鉑金屬錯合物 (cisplatin)。本實驗是利用吡啶釕金屬錯合物會與DNA的鳥糞嘌呤 (guanine)的N7位置鍵結,進而影響DNA的複製及轉錄作用的進行的特性,近一步將不同結構的吡啶釕金屬錯合物加入大腸桿菌的培養中,結果發現吡啶釕金屬錯合物能有效抑制大腸桿菌的生長。萃取出大腸桿菌的蛋白質後,利用蛋白質的等電點及分子量不同將萃取出的蛋白質以等電焦集電泳 (IEF)及SDS-PAGE進行分離,再利用影像比對軟體2-D ImageMaster進行二維凝膠比對,結果發現加入吡啶釕金屬錯合物培養的大腸桿菌在蛋白質表現上的確與未加入吡啶釕金屬錯合物的控制組有所不同,總共發現有43個點有明顯變化。因此,將這43個變化的點切割下來,以胰蛋白酶(trypsin)將蛋白質消化成小片段胜肽,再以奈流液相層析質譜儀(nano LC-MS/MS)進行蛋白質的身分鑑定,進一步分析吡啶釕金屬錯合物所影響的生物功能及代謝路徑。結果發現被吡啶釕金屬所影響的蛋白質表現,多數是被抑制的,少數表現量反而是增多的,這些表現量增多的蛋白質多是能抵抗環境壓力的蛋白質,顯示大腸桿菌的生理功能及代謝路徑被改變,在多數蛋白質被抑制的情形下,可能使大腸桿菌無法執行正常的生理功能及代謝路徑,而導致菌體的死亡。本實驗證實了吡啶釕金屬不但可以抑制大腸桿菌的生長,更會影響大腸桿菌的總體蛋白質表現。藉由了解吡啶釕金屬錯合物所影響蛋白質的代謝路徑及所執行的生理功能,期望能找出吡啶釕金屬錯合物作用在生物體的可能機制,以提供未來設計及發展新藥物的方向。
    Ruthenium complexes have the properties of low systemic toxicity, high water solubility, specificity to antimetastatic tumor, and the potential to displace the anticancer drug of cisplatin. It is interesting to explore the correlation between the structures of ruthenium complexes and the expression of protein. Polypyridyl ruthenium complexes were used to examine this concept. Polypyridyl ruthenium complexes are liable to bind with DNA in N7 position of guanine and lead to the termination of the DNA duplication and transcription. The cell growth was inhibited by using polypyridyl ruthenium complexes. In order to understand the correlation between cell inhibition and protein expression, proteomics was used. The variation of proteins distribution was accomplished by the extraction of E. coli totaprotein with different concentration of polypyridyl ruthenium complexes, the separation of protein in their different pI values and molecular weight by two-dimensional polyacrylamide gel electrophoresis, the analysis by 2-D ImageMaster, the digestion with trypsin, and the detection of the molecular ions with nano LC-MS/MS.
    The result showed that there were 43 spots altered conspicuously. Most protein expression were decreased, and few protein expression were increased. The functions of increased protein expression were the protein which could resist the stress of environment. It revealed that the physical function and metabolic pathway of E. coli was changed, and this reason may cause the death of cell.
    According to the result, the mechanism of polypyridyl ruthenium complexes acting on the cell may provide a consideration in ruthenium-base anticancer drug.
    Appears in Collections:[生命科學研究所] 學位論文

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