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研究生: 周雨駿
論文名稱: 釕金屬錯合物與 DNA 結合之序列選擇性探討
指導教授: 黃文彰
學位類別: 碩士
Master
系所名稱: 化學系
Department of Chemistry
論文出版年: 2007
畢業學年度: 95
論文頁數: 127
中文關鍵詞: 去氧核醣核酸钌金屬錯合物
論文種類: 學術論文
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  • 本論文係探究於 [Ru(bpy)2(dppz)]2+ 及其衍生物 complexs 對特定 AT-rich與 GC-rich DNA 序列之鏡像選擇性與結合性之研究。前人研究指出 [Ru(bpy)2(dppz)]2+ (dppz = dipyrido [3,2-a:2’,3’-c]phenazine) 是個有名的 “light switch” 分子,因為於非質子溶劑中,經照光後可以發出磷光,但是於質子溶劑水溶液中,磷光會被 H+ 給 quench 掉;且還可經由氧化電子傳遞,作為切斷特定 DNA 序列的試劑,此類 complexes中的 dppz ligand,能夠與 DNA 的鹼基產生 π stacking 而插入於 DNA 鹼基對之間,遮蔽掉溶劑中質子與 dppz ligand 上的 N 作用的可能性,於照光後仍可發出磷光。在本實驗中,我們還測試了另三個衍生物[Ru(dmbpy)2(dppz)]2+、[Ru(Cl2bpy)2(dppz)]2+ 和 [Ru(dCOOHbpy)2(dppz)]2+ complexes。在本實驗中我們以上述 Ru complexes 測試了四條雙股右旋且具迴文對稱的 octamers DNA 序列 (d[CGCATGCG])2、(d[CGCTAGCG])2、(d[TATGCATA])2 和 (d[TATCGATA])2。利用 UV/Vis 變溫吸收光譜、UV/Vis 的滴定、circular dichroism (CD) 變溫吸收光譜和 equilibrium dialysis 實驗,來推論出各別 Ru complexes所偏好的 DNA 序列。實驗結果顯示,我們在 [Ru(bpy)2(dppz)]2+、[Ru(dmbpy)2(dppz)]2+ 和 [Ru(Cl2bpy)2(dppz)]2+ 這三者皆可觀察到,其中 [Ru(dmbpy)2(dppz)]2+ 和 [Ru(Cl2bpy)2(dppz)]2+ 最為明顯強烈。且傾向對 AT-rich 的 DNA 序列作結合。然而 DNA 對 [Ru(bpy)2(dppz)]2+ 兩種 enantiomers 的區分能力不是很好,認為應該是對兩種 enantiomers 的結合能力都不差,卻少了專一性。而 [Ru(dCOOHbpy)2(dppz)]2+ 卻是幾乎沒有結合與插入 DNA 鹼基對中的能力。我們也觀察到了各 complexes 對於 DNA 中 purine 和 pyrimidine 彼此交錯排列的序列,有著較強的結合能力,例如 [Ru(bpy)2(dppz)]2+ 是對 (d[CGCATGCG])2,[Ru(dmbpy)2(dppz)]2+ 和 [Ru(Cl2bpy)2(dppz)]2+ 則是對 (d[TATGCATA])2,而 [Ru(dCOOHbpy)2(dppz)]2+ 因結合能力低無能判讀其序列之選擇性。推論於 ancillary ligand 上修飾推拉電子的取代基,是可以有效改善其 enantioselectivity 和與 DNA 結合的能力,且取代基大小比推拉電子性更強烈相關。未來可以以此為基礎,去設計更具專一性和結合能力的 DNA probes 或是 DNA 切割試劑。

    目錄 I 表目錄 IV 圖目錄 V 中文摘要 XI Abstract XIII 第一章 - 1 - 緒論 - 1 - 第一節 過渡金屬錯合物鍵結於 DNA 分子之介紹 - 1 - 第二節 改善插入方式來增加與 DNA 的親和力和辨識力 - 3 - 第三節 DNA 的基本構型 - 8 - 第四節 [Ru(bpy)2(dppz)]2+ 和其衍生物發展 - 13 - 第五節 DNA 的辨認以結構形狀來做選擇 - 16 - 第六節 實驗設計 - 18 - 第二章 - 22 - 實驗部份和原理 - 22 - 第一節 儀器設備 - 22 - 電子吸收光譜 (Electronic absorption spectra) - 22 - 圓二偏極化光光譜 (Circular dichroism spectra) - 22 - 第二節 實驗方法及步驟 - 25 - 樣品取得 - 25 - 樣品製備 (Sample preparation) - 28 - DNA-binding 實驗 - 30 - 第三節 實驗分析原理 - 34 - 圓二偏極化光光譜 (Circular dichroism spectra) 基本原理 - 34 - 分子模擬計算 (Molecular Modeling) 基本原理: - 50 - 第三章 - 51 - 實驗結果與討論 - 51 - 第一節 DNA sequences content 介紹 - 51 - 第二節 電子吸收光譜變溫實驗 (UV/Vis Termal Denaturation Experiment) - 55 - 第三節 電子吸收光譜滴定實驗 (Electronic absorption spectral titration) - 59 - 第四節 Equilibrium Dialysis Experiment –Binding % - 71 - 第五節 Equilibrium Dialysis Experiment - one Ru complex to four kinds of DNA sequences - 76 - 第六節 Equilibrium Dialysis Experiment - one DNA sequence to four kinds of Ru complexes - 92 - 第七節 CD Termal denaturation experiment - 101 - [Ru(bpy)2(dppz)]2+ : DNA -1 : 1 - 102 - [Ru(dmbpy)2(dppz)]2+ : DNA -1 : 1 - 108 - [Ru(Cl2bpy)2(dppz)]2+ : DNA -1 : 1 - 115 - [Ru(dCOOHbpy)2(dppz)]2+ : DNA -1 : 1 - 120 - 第四章 - 125 - 結論 - 125 - 參考文獻 - 127 -

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