臺灣博碩士論文加值系統

單股DNA結合蛋白質(SSB)在DNA之代謝中，包括複製、修復及重組的過程中扮演重要的角色。SSB之N端區域形成一種寡醣核苷酸結合區使之能與DNA結合，而C端是一段具有高度保留特性的酸性胺基酸序列，其重要性為與其他參與DNA代謝的蛋白質作用。為了解單股DNA與SSB在溶液中的結合情形，我們利用螢光滴定法與凝膠阻滯分析法觀察其複合物的形成，其結合方式為一個SSB(其在生理環境下作用為四聚體)與25到30個核苷酸結合。本實驗室利用X光晶體繞射實驗決定C端去尾之胃幽門螺旋桿菌SSB (HpSSBc)之結構，所得的晶體最高解析度達2.3 Å。在晶體中顯示出兩條單股聚核苷酸(長度各為35個胸腺嘧啶核苷酸)與一個SSB四聚體結合。單股DNA藉由靜電力與疏水性堆疊作用力與HpSSBc纏繞。HpSSBc四聚體在水溶液中以30個核苷酸之結合模式作用，而我們在晶體中則觀察到65個核苷酸之結合模式。這兩種不同的結合模式可能分別作用在DNA複製、修復及重組的過程中。

Contents
中文摘要
Abstract
Contents
Chapter 1 Introduction
1.1 Helicobacter pylori
1.2 Single-stranded DNA binding protein (SSB)
1.3 The binding modes of SSB
1.4 The C-terminal sequence of SSB
1.5 The association proteins of SSB
1.6 The characteristics of other SSBs
Chapter 2 Material and methods
2.1 Cloning , protein expression and purification
2.2 Analytical ultracentrifugation
2.3 Analytical gel filtration chromatography
2.4 Fluorescence titration
2.5 Gel shift assay
Chapter 3 Results
3.1 Protein purification and identification
3.1.1 HpSSB
3.1.2 C-terminal truncated HpSSB (HpSSBc)
3.2 HpSSB and HpSSBc binding mode determination
3.3 Overall structure of HpSSBc
Chapter 4 Discussion
4.1 Comparison of ssDNA binding site size between full-length HpSSB and HpSSBc
4.2 The binding mode of HpSSBc
4.3 Comparison of ssDNA binding mode between SSBs
Appendix Figures and Tables
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