一种简便快速工艺制备重组人胸腺肽α_1  

A simple and rapid process for preparing rhthymosin α_1 with transgenic Escherichia coli

作　　者：秦伟涛[1] 陈劲春[1] 李冬冬[1] 

机构地区：[1]北京化工大学生命科学与技术学院,北京100029

出　　处：《北京化工大学学报（自然科学版）》2007年第3期296-299,共4页Journal of Beijing University of Chemical Technology(Natural Science Edition)

摘　　要：将构建的含有胸腺肽α1融合蛋白的编码基因插入表达质粒pET28a(+)中,并在大肠杆菌BL21中进行表达,该菌经IPTG诱导融合蛋白高表达后,离心收集菌体,超声波破碎,包含体裂解,目标蛋白质的体外变性复性后,经DEAE-Sepharose Fast Flow阴离子交换层析纯化,每升菌液中可获得260 mg融合蛋白。经重组肠激酶酶切反应、Ni-Sepharose亲和层析,Sephadex G-25柱纯化后,得到了纯度达97%的重组人胸腺肽α1,该纯化工艺简单快速,经三步层析即可得到纯度达97%的重组人胸腺肽α1,且产量达82 mg/L,为大批量用细菌表达重组人胸腺肽α1及纯化提供了参考。Human thymosin α1, a small hormone with a length of 28 amino acids, is a powerful antiviral drug for treatment of HBV, SARS and various cancers. After consideration of the feasibility of producing human thymosin α1 on a large scale, an over-expression vector has been constructed based on the plasmid pET28a（ ＋ ）. After induction by isopropyl β-D-1-thiogalactopyranoside （IPTG）, the observed yield of crude fusion proteins was up to 260mg/L. In order to obtain the purified target proteins, three different chromatographic columns, namely DEAE-Sepharose FF, Ni-chelating-Sepharose and Sephadex G-25, were operated in series. The purity of the resulting rhThymosin at was more than 97%, as shown by RP-HPLC analysis. Finally, 82 mg of purified rhThymosin α1 was obtained from α1 liter culture.

关 键 词：重组人胸腺肽α1 大肠杆菌 Ni—Sepharose亲和层析 融合蛋白 

分 类 号：Q512.7[生物学—生物化学]