| 水稻谷胱甘肽磷脂氢过氧化物酶的表达、纯化及晶体生长条件初筛 |
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| 引用本文： | 王丰,刘进元. 水稻谷胱甘肽磷脂氢过氧化物酶的表达、纯化及晶体生长条件初筛[J]. 高等学校化学学报, 2007, 28(9): 1701-1706 |
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| 作者姓名： | 王丰 刘进元 |
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| 作者单位： | 清华大学生物科学与技术系分子生物学实验室,教育部蛋白质科学重点实验室,北京,100084;清华大学生物科学与技术系分子生物学实验室,教育部蛋白质科学重点实验室,北京,100084 |
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| 基金项目： | 国家高技术研究发展计划(863计划) , 国家重点基础研究发展计划(973计划) , 国家自然科学基金 , 清华-裕元医学科学研究基金 |
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| 摘 要： | 将水稻PHGPx(OsPHGPx)的编码序列克隆到表达载体pGEX-6P-1上, 并转化为大肠杆菌进行表达. 通过GST亲和层析、离子交换层析和凝胶过滤层析, 制备了可用于晶体学研究的OsPHGPx, 其纯度超过95%, 具备明显的PHGPx活性. 质谱显示OsPHGPx的精确分子量为19642.5553, 与理论分子量基本一致. OsPHGPx在多个晶体生长条件下出现微晶. 三维结构同源建模显示 OsPHGPx的结构为硫氧还蛋白折叠形式.
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| 关 键 词： | 水稻 谷胱甘肽磷脂氢过氧化物酶 表达 纯化 晶体生长 三维结构建模 |
| 文章编号： | 0251-0790（2007）09-1701-06 |
| 收稿时间： | 2006-11-21 |
| 修稿时间： | 2006-11-21 |
Expression, Purification and Crystal Growing Conditions of Recombinant Oryza sativa PHGPx |
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| WANG Feng,LIU Jin-Yuan. Expression, Purification and Crystal Growing Conditions of Recombinant Oryza sativa PHGPx[J]. Chemical Research In Chinese Universities, 2007, 28(9): 1701-1706 |
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| Authors: | WANG Feng LIU Jin-Yuan |
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| Affiliation: | Laboratory of Molecular Biology and Protein Science Laboratory of the Ministry of Education, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China |
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| Abstract: | Phospholipid hydroperoxide glutathione peroxidase(PHGPx)is a unique antioxidant enzyme that directly reduces lipid hydroperoxides in biomembranes.It plays potential important roles in oxidative stress response.Oryza sativaPHGPx gene was cloned into expression vector pGEX-6P-1 and transformed into E.coli strain BL21(DE3).OsPHGPx for crystals was prepared with employing Glutathione SepharoseTM affinity,cation-exchange and gel filtration chromatography.The purity of the purified OsPHGPx was over 95%.OsPHGPx showed an obvious PHGPx activity towards lipid hydroperoxides.MALDI-TOF analysis shows that the exact molecular weight of OsPHGPx was 19275.568,which was in accordance with the theoretical molecular weight.The microcrystals of OsPHGPx were obtained under several conditions.In addition,a tertiary structure model of the OsPHGPx generated from http://swissmodel.expasv.org/displayed the thioredoxin fold. |
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| Keywords: | Oryza sativa Phospholipid hydroperoxide glutathione peroxidase Expression Purification Microcrystal growth of OSPHGPx 3D structure modeling |
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