|本期目录/Table of Contents|

[1]石刚,闫娟,岳昌武,等.人α-防御素HNP-2基因在大肠杆菌中的表达[J].应用与环境生物学报,2008,14(01):78-082.
 SHI Gang,et al..Expression of Human αdefensin HNP2 Gene in Escherichia coli[J].Chinese Journal of Applied & Environmental Biology,2008,14(01):78-082.
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人α-防御素HNP-2基因在大肠杆菌中的表达()
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《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
14卷
期数:
2008年01期
页码:
78-082
栏目:
综述
出版日期:
2008-01-15

文章信息/Info

Title:
Expression of Human αdefensin HNP2 Gene in Escherichia coli
作者:
石刚闫娟岳昌武戚天胜陈守春张义正
(1四川大学生命科学学院成都610064; 2成都地奥制药集团有限公司成都610041)
Author(s):
SHI Gang et al.
(1College of Life Sciences, Sichuan University, Chengdu 610064, China) (2Chengdu Di’ao Pharmaceutical Company, Chengdu 610041, China)
关键词:
关键词人α防御素基因表达大肠杆菌环化酰胺化
Keywords:
Keywords human αdefensin gene expression Escherichia coli cyclization amidation
摘要:
摘要 通过半化学合成方法获得具有大肠杆菌密码子偏爱性的人α防御素(Human αdefensin或human neutrophil peptide, HNP)HNP2基因.将其克隆到pET32a(+)表达载体,构建了重组表达质粒pET32a(+)/HNP2.分别转化大肠杆菌BL21(DE3), ER2566以及Origami B(DE3)宿主菌,经过表达条件的优化,结果在大肠杆菌Origami B(DE3)宿主菌中得到了HNP2基因高效率可溶性的表达,融合蛋白表达量占细菌总蛋白的60%以上.为避免表达产物水解,得到更稳定的人α防御素,尝试以环化形式和酰胺化形式表达人α防御素.为得到环化形式的表达,采用intein作为工具,利用其可剪接extein的特点,将其N端区域和C端区域分别融合到HNP2的C端和N端,其两端的intein片段可将其拼接成环肽.以此为基础构建了环化表达质粒,并对表达条件进行了探索.本文采用了一种新的酰胺化方法,即将intein与HNP2融合表达,intein对目的短肽进行酰胺化修饰.图6表1参16
Abstract:
Abstract A hemichemical synthesis method was adopted to get HNP2 gene which had code bias in Escherichia coli. The HNP2 gene was inserted into expression vector pET32a(+), and a expression plasmid pET32a(+)/HNP2 was constructed. After the plasmid was transformed into E. coli BL21 (DE3), ER2566 and Origami B(DE3), respectively, and the expression conditions were optimized, high level soluble expression of HNP2 gene was acquired in the host bacteria Origami B(DE3). The soluble fusion protein accounted for 60% of the total bacterial protein. To avoid the hydrolyzation of the expressed product by the cellular proteases and obtain more stable human αdefensin, it was attempted to express HNP2 gene in cyclic or amidation form. To get the cyclic peptide of defensin, the Nterminal and Cterminal intein fragments were fused to Cterminal and N terminal target peptide, respectively. So the target peptide could be cyclized by the two fused intein fragments. The recombinant expression plasmid was constructed and the cyclic HNP protein might be obtained. A new method was adopted to study the amidation of HNP expressed as fused protein. The recombinant expression plasmids were constructed by fusing the HNPcoding fragment with intein and the amidation of HNP2 might be observed. Fig 6, Tab 1, Ref 16

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更新日期/Last Update: 2008-03-05