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蛋白酪氨酸磷酸酶PTP1B催化活性区的原核表达及活性分析
Expression and Activity Analysis of Catalytic Domain of PTP1B

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蛋白酪氨酸磷酸酶PTP1B催化活性区的原核表达及活性分析.pdf (616.9Kb)
Date
2008-04-25
Author
王生育
颜江华
潘阳霖
李学军
陈忠
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  • 医学院-已发表论文 [2567]
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Abstract
从GenBank获得人PTP1B催化活性区(PTP1Bc)氨基酸序列(1~301aa),通过重叠PCR获得PTP1Bc基因。构建pET-22b(+)/PTP1Bc原核表达载体,转化大肠杆菌BL21(DE3),阳性重组子IPTG诱导表达,Ni柱纯化蛋白。目的蛋白以包涵体的形式表达,表达量占菌体总蛋白30%以上。纯化后,蛋白纯度达95%以上。Western blotting结果表明所得的蛋白可与抗PTP1B抗体发生特异性结合;酶活实验证实复性的蛋白具有一定的磷酸酶活性。PTP1Bc基因的构建、表达纯化及活性分析,为进一步的功能研究奠定了基础。 【英文摘要】 The amino acid sequence (1-301aa) coding the human PTP1B catalytic domain (PTP1Bc) was obtained from the GenBank.The PTP1Bc gene was constructed by overlapping PCR,then was inserted into vector pET-22b(+) and expressed efficiently in E.coli BL21(DE3) under optimum condition after IPTG induction.The proteins were expressed mainly as inclusion bodies with the yield of more than 30% of total bacterial proteins.The expressed products were purified through Ni~(2+)-affinity chromatographic column.After purificati...
Citation
生物工程学报 Chinese Journal of Biotechnology,200824(4):553-557
URI
https://dspace.xmu.edu.cn/handle/2288/8488

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