[1]李国强,周亮,李安国,等.斑节对虾carcininPm3的表达及其无标签多肽的获得[J].深圳大学学报理工版,2020,37(1):9-16.[doi:10.3724/SP.J.1249.2020.01009]
 LI Guoqiang,ZHOU Liang,LI Anguo,et al.Expression of tag-free carcininPm3 of Penaeus monodon[J].Journal of Shenzhen University Science and Engineering,2020,37(1):9-16.[doi:10.3724/SP.J.1249.2020.01009]
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斑节对虾carcininPm3的表达及其无标签多肽的获得()
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《深圳大学学报理工版》[ISSN:1000-2618/CN:44-1401/N]

卷:
第37卷
期数:
2020年第1期
页码:
9-16
栏目:
生物工程
出版日期:
2020-01-08

文章信息/Info

Title:
Expression of tag-free carcininPm3 of Penaeus monodon
文章编号:
202001002
作者:
李国强周亮李安国王潮岗
深圳大学生命与海洋科学学院,深圳市海洋生物资源与生态环境重点实验室,广东省海洋藻类开发与应用工程重点实验室,广东深圳 518060
Author(s):
LI Guoqiang ZHOU Liang LI Anguo and WANG Chaogang
College of Life Sciences and Oceanography, Shenzhen Key Laboratory of Marine Bioresources and Ecology, Guangdong Technology Research Center for Marine Algal Bioengineering, Shenzhen University, Shenzhen 518060, Guangdong Province, P.R.China
关键词:
基因工程斑节对虾抗菌肽crustin家族carcininPm3基因原核表达重组蛋白亲和层析
Keywords:
genetic engineering Penaeus monodon antimicrobial peptide family crustin carcininPm3 prokaryotic expression recombination protein affinity chromatography
DOI:
10.3724/SP.J.1249.2020.01009
文献标志码:
A
摘要:
crustin是对虾中重要的抗菌肽,本研究获得的carcininPm3属于斑节对虾crustin家族,是一种新型抗菌肽,成熟肽含有92个氨基酸,与其他对虾抗菌肽的同源性小于45%.根据其氨基酸序列和大肠杆菌(Escherichia coli, E. coli)的密码子偏好性设计carcininPm3的基因序列,将carcininPm3连接到原核表达载体pSmartI-SUMO上,构建重组表达载体pSmartI-SUMO-carcininPm3, 再转化至原核表达菌株E. coli BL21(DE3)中,通过异丙基-β-D-硫代半乳糖苷(isopropyl β-D-thiogalactopyranoside,IPTG)诱导SUMO-carcininPm3的融合表达,进一步优化其表达条件,获得表达量大、可溶性高的重组蛋白.通过亲和层析纯化得到重组蛋白,蛋白的质量浓度达到195 μg/mL.利用SUMO酶去除SUMO融合标签,获得没有外源标签的carcininPm3抗菌肽,可用于蛋白功能分析、抗体制备等研究.研究结果为carcininPm3功能的分析及应用提供参考.
Abstract:
Crustins are one of the vital antimicrobial peptides in shrimp. CarcininPm3, as one of the members of the crustin family, contained 92 amino acids in its mature peptide. It is a novel antimicrobial peptide in shrimp because of only 45% homology to other species. In this paper, firstly, sequence of carcininPm3 was redesigned according to its mature peptide sequence and codon bias of Escherichia coli (E. coli), and then inserted into pSmartI-SUMO vector to finally obtain pSmartI-SUMO-carcininPm3 recombinant plasmid. This plasmid was introduced into E. coli BL21 (DE3) and the transformants were capable to express SUMO-carcininPm3 in fusion form when induced by IPTG. Further studies were proceeded to optimize the expression of SUMO-carcininPm3, acquiring recombinant protein with large quantity and high solubility. Finally, this recombinant protein was purified by affinity chromatography and the content was up to 195 μg/mL. After treated with SUMO protease, the SUMO tag was removed and tag-free carcininPm3 peptide was obtained. The above results provid a foundation to the functional analysis and application of carcininPm3.

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备注/Memo

备注/Memo:
Received:2018-11-28;Accepted:2019-01-23
Foundation:National Natural Science Foundation of China (31470389); Shenzhen Science and Technology Plan (JCYJ20160422171614147, JCYJ20170818101523761, JCYJ20170302144605664)
Corresponding author:Associated professor WANG Chaogang. E-mail: charlesw@szu.edu.cn
Citation:LI Guoqiang,ZHOU Liang,LI Anguo, et al. Expression of tag-free carcininPm3 of Penaeus monodon[J]. Journal of Shenzhen University Science and Engineering, 2020, 37(1): 9-16.(in Chinese)
基金项目:国家自然科学基金资助项目(31470389);深圳市科技计划资助项目(JCYJ20160422171614147, JCYJ20170818 101523761, JCYJ20170302144605664)
作者简介:李国强(1993—)男,深圳大学硕士研究生.研究方向:水生生物学.E-mail:504493562@qq.com
引文:李国强,周亮,李安国,等.斑节对虾carcininPm3的表达及其无标签多肽的获得[J]. 深圳大学学报理工版,2020,37(1):9-16.
更新日期/Last Update: 2020-01-30