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Hyper-production of porcine contagious pleuropneumonia subunit vaccine proteins in Escherichia coli by developing a bicistronic T7 expression system

文献类型： 外文期刊

作者： Sun, Manman 1 ; Gao, Alex Xiong 5 ; Li, An 1 ; Ledesma-Amaro, Rodrigo 2 ; Wang, Peng 4 ; Chen, Wenchao 6 ; Bai, Zhonghu 1 ; Liu, Xiuxia 1 ;

作者机构： 1.Jiangnan Univ, Natl Engn Res Ctr Cereal Fermentat & Food Biomfg, Wuxi 214112, Peoples R China

2.Imperial Coll London, Dept Bioengn, London, England

3.Imperial Coll London, Imperial Coll Ctr Synthet Biol, London, England

4.Chinese Acad Sci, Hefei Inst Phys Sci, Key Lab High Magnet Field & Ion Beam Phys Biol, Hefei, Peoples R China

5.Hong Kong Univ Sci & Technol, Div Life Sci, Hong Kong, Peoples R China

6.Chinese Acad Agr Sci, Oil Crops Res Inst, Wuhan, Hubei, Peoples R China

关键词： bicistronic T7 expression system; Escherichia coli; hyper-production; PCP subunit vaccine protein; recombinant expression

期刊名称：BIOTECHNOLOGY JOURNAL （ 影响因子：4.7； 五年影响因子：4.5 ）

ISSN： 1860-6768

年卷期： 2024 年 19 卷 1 期

页码：

收录情况： SCI

摘要： The ApxII toxin and the outer membrane lipoprotein (Oml) of Actinobacillus pleuropneumoniae are important vaccine antigens against porcine contagious pleuropneumonia (PCP), a prevalent infectious disease affecting the swine industry worldwide. Previous studies have reported the recombinant expression of ApxII and Oml in Escherichia coli; however, their yields were not satisfactory. Here, we aimed to enhance the production of ApxII and Oml by constructing a bicistronic expression system based on the widely used T7 promoter. To create efficient T7 bicistronic expression cassettes, 16 different fore-cistron sequences were introduced downstream of the T7 promoter. The expression of three vaccine antigens Oml1, Oml7, and ApxII in the four strongest bicistronic vectors were enhanced compared to the monocistronic control. Further optimization of the fermentation conditions in micro-well plates (MWP) led to improved production. Finally, the production yields reached unprecedented levels of 2.43 g L-1 of Oml1, 2.59 g L-1 of Oml7, and 1.21 g L-1 of ApxII, in a 5 L bioreactor. These three antigens also demonstrated well-protective immunity against A. pleuropneumoniae infection. In conclusion, this study establishes an efficient bicistronic T7 expression system that can be used to express recombinant proteins in E. coli and achieves the hyper-production of PCP vaccine proteins.

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