臺灣博碩士論文加值系統

活細胞無時無刻都受到外在環境及自源性的氧化物質或自由基等應力而造成DNA的鹼基被氧化、分解，以致於使得基因產生變異。G*(8-oxodG)是dG被氧化的衍生物，亦為生物體內最常見的氧化性鹼基之一。近年來G*已被廣泛的研究，甚至被當成一些慢性病或是老化程度的指標。本實驗的研究重點在於G*鹼基在髮夾式DNA構造的影響，以dC及dA配對之構型來加以探討，研究其結構特異性。
在實驗的結果上，我們得到下列結論：G*採反式醣甘鍵角(anti glycosidic torsion angle)與反式(anti form)的dC形成配對；採用順式醣甘鍵角(syn glycosidic torsion angle)與anti form的dA形成配對。雖然由熱力學的證據顯示，此種錯誤配對仍有很高的穩定性(G:C Tm：78°C；G*:C Tm：70°C；G*:A Tm：66°C)，但由1D NOE及H2O-NOESY實驗的光譜可清楚的看出，由於G*的併入，在髮夾式雙股的結構中所產生的立體障礙效應(steric effect)，造成其穩定性改變，另外我們也需要更進一步的實驗來證明此種改變是使得DNA聚合酵素(DNA polymerase)無法修補這樣不正常配對，而需要Mut蛋白質來進行此動作的原因。

Living cells always incur stress such as oxidative substance from environment or intrinsic free radical to result in oxidation or degradation of the base of DNAs, making the gene to have an alternation as a consequence. The G*(8-oxodG), one of the common oxidative bases in vivo, is an oxidative derivative from dG. G* has been extensively explored lately to be used as a marker of chronic diseases or the stage of aging. The central effort of this research is to determine the global influence of the structure of hairpin DNAs containing G* to having a base pair with dA or dC, respectively.
The G* matched with dC adapted anti glycosidic conformation while the dC is in anti form. However, G* had syn glycosidic torsion angle when it paired with dA. Though these abnormal base pairs mentioned above having high stability from thermodynamic evidences(G:C Tm：78°C；G*:C Tm：70°C；G*:A Tm：66°C), the oxidation of dG to G* induces the steric effect, observed by the 1D NOE and H2O-NOESY, in the motif of double stranded hairpin structure. This unfavorable effect leads to a destabilizing impact on hairpin DNAs. Further research is needed to having a picture on how these errors to be repaired by Mut proteins rather than DNA polymerase.

圖目錄
表目錄
中文摘要……………………………………………………1
英文摘要……………………………………………………2
壹、前言
一、8-OxodG簡介…………………………………………4
二、合成DNA序列及構型之探討…………………………7
三、研究目標………………………………………………8
貳、基本原理
一、核酸介紹………………………………………………9
二、核磁共振基本原理……………………………………12
三、NOE效應……………………………………………….13
參、實驗方法及步驟
一、 藥品及儀器……………………………………….14
二、 DNA之合成………………………………….16
三、 光譜資料的收集……………………………18
四、 數據處理及光譜分析………………………19
肆、結果與討論
一、一維核磁共振光譜………………………………22
(1)、一維核磁共振變溫實驗……………………25
(2)、一維核磁共振光譜的分析(1D-NOE)………29
二、確認非交換性的氫………………………………37
(1)、OtA序列每個region的循序判定結果….40
(2)、OtA序列中心損傷部位(central
lesion site)的特異性分析…………………….40
(3)、TTTT圈環結構的定性描述…………………44
伍、結論……………………………………………………45
陸、附錄……………………………………………………48
柒、參考文獻………………………………………………51

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許財誠(1998)含有8-oxodG鹼基之髮夾式去氧核醣核酸其熱力學的 分析 Thermodynamic Stability of Hairpin DNAs Containing an 8-Oxodeoxyguanosine