| 杨诏旭夏宁金成窦科峰.幽门螺杆菌PPK 的纯化与功能分析*[J].现代生物医学进展英文版,2012,12(6):1017-1020. |
| 幽门螺杆菌PPK 的纯化与功能分析* |
| The Purification and Function Analysis of Helicobacter PyloriPolyphosphate Kinase |
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| DOI： |
| 中文关键词: 幽门螺杆菌 多聚磷酸激酶 多聚磷酸盐 |
| 英文关键词: Helicobacter pylori Polyphosphate kinase Polyphosphate |
| 基金项目:国家自然科学基金项目（81000164） |
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| 中文摘要: |
| 目的：表达纯化幽门螺杆菌多聚磷酸激酶，并测定其功能。方法：将幽门螺杆菌多聚磷酸激酶基因克隆入原核表达载体PQE
80L 中，在大肠杆菌(E.coli) DH5-α 中表达。用BD Talon resin 纯化目的蛋白。并在体外测定其合成多聚磷酸盐及转化多聚磷酸盐
至ATP 的能力。结果：成功构建了原核表达载体，得到高表达量的融合蛋白。经BD Talon resin 纯化获得较高纯度的His- 多聚磷
酸激酶N 端融合蛋白。体外实验证实该酶可以有效合成不同链长的多聚磷酸盐，并且在适当条件下可将多聚磷酸盐转化为ATP。
结论：利用原核表达载体可很好表达幽门螺杆菌多聚磷酸激酶，纯化后的蛋白具有良好生物活性，是一个具备合成多聚磷酸盐及
转化其为ATP 的双向功能的酶。 |
| 英文摘要: |
| Objective: To express and purify the polyphosphate kinase of Helicobacter pylori and analyze its function in vitro.
Methods: The polyphosphate kinase gene of Helicobacter pylori were inserted into prokaryotic expression vector PQE 80L. The
expression of the gene was induced in E.coli DH5-α. Then the 6-His fused protein was purified by using BD Talon resin. The activity of
synthesizing polyphosphate and transfering polyphosphate to ATP of PPK were detected. Results: The expression of PPK was
constructed and His-fused PPK was induced. The N-His fused protein was purified by BD Talon resin. This protein can synthesize
Polyphosphate of different length or transfer polyphosphate to ATP under certain condition. Conclusions: PPK had good expression in
prokaryotic expression vector PQE 80L and the protein showed biological activity of synthesizing polyphosphate and transferring
polyphosphate to ATP. |
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