Par系統在染色質分離、啟動孢子形成和對細胞週期的進行扮演重要的角色。ParABS系統由ParA、ParB兩個蛋白質和parS序列所組成。在胃幽門螺旋桿菌中， HpSoj(Hp1139)及HpSpo0J (Hp1138)這兩個蛋白質分別具有ParA及ParB的功能。HpSoj除了是一個ATPase之外，有關HpSoj的其他功能都還沒有被證實，例如在HpSoj的結構中，HpSoj如何與核酸結合和HpSoj如何與HpSpo0J作用是值得研究的。HpSoj經由大量表現並純化得到分子量約為29.2kDa。利用膠體過濾的實驗發現不論有沒有ATP, HpSoj 主要是以單體形態存在。而在電泳遲滯的實驗中，結果指出有ATP時，HpSoj除了對質體pUC19有較強的核酸結合能力，與小片段的parS DNA 也有結合的能力。我們以PEG3350為主沉澱劑培養得到HpSoj-ATP複合物的晶體，其X光繞射的解析度為1.9 Å，此晶體的空間群(space group)為P212121，晶格參數為a=48.1 Å, b=93.3 Å, c=110.9 Å，不對稱單元中具有兩個分子。我們嘗試利用HpSoj 的同源蛋白進行分子置換法(molecular replacement)計算HpSoj的結構相位角，以求得完整的立體結構，藉此了解HpSoj與DNA和Spo0J結合的位置，進而得知HpSoj的生物功能與其作用機制。

The partition system played important roles in segregation of chromosome, initiation of sporulation and progression of cell cycle. The ParABS system consisted of two proteins, ParA and ParB, and a parS sequence. In Helicobacter pylori, HpSoj (Hp1139) and HpSpo0J (Hp1138) was putative ParA and ParB proteins, respectively. Except HpSoj is an ATPase; there are few data to support other information. For example how Soj binds DNA and how Soj interacts with HpSpo0J in the HpSoj structure is worth investigating. HpSoj overexpressed and purified with a molecular weight of 29.2 kDa. Whether ATP is present or not, HpSoj was mainly a monomer by size exclusion chromatography. In Electrophoretic mobility shift assay showed that DNA binding activity of ATP-bound HpSoj with pUC19 was stronger than HpSoj. Furthermore, ATP-bound HpSoj had parS-binding activity. The HpSoj-ATP complex crystal was grown using PEG 3350 as a precipitant. Crystal structure of the HpSoj-ATP was determined at 1.9 Å. The HpSoj-ATP complex crystal belongs to P212121 space group with unit cell parameters a=48.1 Å, b=93.3 Å, c=110.9 Å, containing two molecules per asymmetry unit. We try to use the homologous protein as a search model to do the phase calculation by molecular replacement method, and the structure phase determination of HpSoj is still continuing. The three dimensional structure of HpSoj will help us to understand biological function and catalytic mechanisms of HpSoj.

中文摘要 Ⅱ
Abstract Ⅲ
謝誌 Ⅳ
Contents Ⅴ
Chapter 1 Introduction
1-1 Helicobacter pylori 1
1-2 The plasmid partitioning system 2
1-3 The chromosomally-encoded Par systems 3
1-4 The ParABS system 4
1-5 Background of Soj (ParA) 4

Chapter2 Materials and Methods
2-1 Bacterial strain, plasmid, enzyme, and kit reagent 6
2-2 Cloning of HpSoj 6
2-3 Protein Expression and Purification 7
2-4 Size Exclusion Chromatography 8
2-5 Agarose Electrophoretic Mobility Shift Assay 8
2-6 Acrylamide Electrophoretic Mobility Shift Assay 9
2-7 Crystallization 9
2-8 Data Collection and Space Group Determination 10
2-9 Self-Rotation Functions 11

Chapter3 Results and Discussion
3-1 Bioinformatics of HpSoj 12
3-2 Purification and Characterization of HpSoj 12
3-3 Size Exclusion Chromatography 13
3-4 DNA binding activity of HpSoj 14
3-5 Crystallization 15
3-6 Data Collection and Space Group Determination 16
3-7 Self-Rotation Functions 17

Chapter4 Conclusion 18
Figures 20
Tables 33
Reference 39

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