High lever soluble expression and purification of recombinant human IL-18 in E.coli

PENG Yan; et al

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High lever soluble expression and purification of recombinant human IL-18 in E.coli

更新时间：2024-01-17

- High lever soluble expression and purification of recombinant human IL-18 in E.coli
- Journal of Chongqing Medical University Issue 4, Pages: 524-527(2006)
- 作者机构：
  
  1. 重庆医科大学生物化学与分子药理重点实验室
  2. 重庆医科大学生物化学与分子药理重点实验室,重庆,400016
- 作者简介：
- 基金信息：
- DOI：
  CLC： R392
- Published：2006，
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[1]彭彦,王勇,宋方洲,王亚平.IL-18在大肠杆菌中的可溶性表达和纯化[J].重庆医科大学学报,2006(04):524-527+531.

PENG Yan, et al. High lever soluble expression and purification of recombinant human IL-18 in E.coli. [J]. Journal of Chongqing Medical University (4):524-527(2006)
[1]彭彦,王勇,宋方洲,王亚平.IL-18在大肠杆菌中的可溶性表达和纯化[J].重庆医科大学学报,2006(04):524-527+531. DOI：

PENG Yan, et al. High lever soluble expression and purification of recombinant human IL-18 in E.coli. [J]. Journal of Chongqing Medical University (4):524-527(2006) DOI：

摘要

目的:研究人IL-18成熟蛋白（mhIL-18）在大肠杆菌中的高效的非包涵体表达以及纯化方案。方法:利用PCR方法扩增出mhIL-18基因

构建融合型表达载体pPTYB1-IL18

转化入大肠杆菌ER2566中

IPTG诱导表达

优化表达条件

超声裂解细胞

采用SDS-PAGE和Western blot分析重组蛋白的表达

亲和层析后用MTT法检测表达mhIL-18的活性。结果:成功构建载体并转化入ER2566后

经IPTG诱导

表达量可占菌体总蛋白的26%以上

其中80%以上的mhIL-18融合蛋白以可溶、有活性的形式存在。亲和层析后的mhIL-18纯度可达95%

具有显著的IL-18的生物学活性。结论:成功的在大肠杆菌中的高效以非包涵体表达mhIL-18

并具有显著的IL-18的生物学活性。

Abstract

Objective: Objective To study high-level expression and purification of recombinant mature human IL-18(mhIL-18)in E.coli with soluble form.Methods: The gene of mhIL-18 was amplified with PCR.Then

the recombinant expression plasmid pTYB1IL-18 was constructed by cloning mhIL-18 into pTYB1 vector and transformed to E.coli ER2566.IPTG was added into the E.coli ER2566 culture to express recombinant fusion protein.After the harvested bacteria dispersed with ultrasound

SDS-PAGE and Western blot were used to analyze the expression of recombinant fusion protein.The activity of mhIL-18 which had been purified by affinity chromatography was tested by MTT.Results:The cloned sequence of mhIL-18 was identical with the sequence in GenBank.After the induction by IPTG inducing

the fusion protein of mhIL-18 covered over 26% total bacterial proteins

which included 80% active fusion protein.Purification of recombinant expressed mhIL-18 purity was 95%

which had the activity of the mhIL-18.Conclusion: High level expression and purification of recombinant human IL-18 is succeeded which has the activity.

关键词

白介素-18原核表达纯化

Keywords

Interleukin-18Prokaryoticexpressionpurification

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