| CD59融合蛋白表达载体构建及其在大肠杆菌中的表达 |
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| 引用本文： | 周镜然,白云,姜曼,黎万玲. CD59融合蛋白表达载体构建及其在大肠杆菌中的表达[J]. 免疫学杂志, 2000, 16(2): 96-98 |
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| 作者姓名： | 周镜然 白云 姜曼 黎万玲 |
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| 作者单位： | 第三军医大学免疫学教研室,重庆,400038 |
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| 基金项目： | 国家自然科学基金资助项目(39770163) |
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| 摘 要： | 探索ＧＰＩ－锚固蛋白的体外表达，获得具有生物学活性的可溶性游离ＣＤ５９分子。方法通过ＰＣＲ选择性扩增编码成熟ＣＤ５９氨基酸序列的基因片段，并将之克隆入ＰｉｎＰｏｉｎｔＸａ－３质粒中。ＩＰＴＧ诱导融合蛋白表达，产物经亲和素树脂亲和层析。在丛外微量反应性溶血抑制实验中检测重组ＣＤ５９纯化样品的生物学活性。
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| 关 键 词： | CD59 融合蛋白 大肠杆菌 基因表达 |
| 文章编号： | 1000—8861(9900)02—0096—03 |
| 修稿时间： | 1999-10-06 |
Recombination and expression of soluble human CD59 in E. Coli |
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| ZHOU Jing-ran,BAI Yun,JIANG Man,LI Wan-ling. Recombination and expression of soluble human CD59 in E. Coli[J]. Immunological Journal, 2000, 16(2): 96-98 |
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| Authors: | ZHOU Jing-ran BAI Yun JIANG Man LI Wan-ling |
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| Affiliation: | ZHOU Jing-ran,BAI Yun,JIANG Man,LI Wan-ling;(Department of Immunology, the Third Military Medical University, Chongqing 400038,China) |
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| Abstract: | Objective To obtain soluble recombinant human CD59 with high activity. Methods The gene sequence which codes natural CD59 extracellular portion containing 77 amino acids was amplified with PCR. The fragment was cloned into prokaryotic expression vector PinPoint Xa-3. The recombinant protein expression was induced by IPTG and biotinylated products were purified by avidin resin. The activity of purified recombinant CD59 was evaluated by reactive assay in vitro. Resuits In PinPoint Xa-3 expression system, we obtained recombinant CD59 with a molecular wright of 22kDa. The thin layer gel scanning analysis showed that about 7% of total cell protein is newly expressed fusion protein. SDS-PAGE assay showed that the product affinity-purified by avidin resin has high purity. The reactive assay revealed that the purified protein had the activity of inhibiting hemolysis. Conclusion The fusion protein has the activity of natural CD59 to some extent. |
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| Keywords: | recombinant CD59 fusion protein expression |
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