Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus
Intein-mediated expression and purification of common carp IFN-γ and its protective effect against spring viremia of carp virus
复制标题
内含肽介导的鲤鱼IFN-γ表达纯化及其对鲤鱼春季病毒血症的保护作用
DOI：
10.1016/j.fsi.2019.03.022
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发表时间：
2019-05-01
影响因子：
4.7
通讯作者：
Liu, Yi
中科院分区：
文献类型：
--
作者：
Fu, Jianping;Yi, Zhiqiang;Liu, Yi
IFN-gamma is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-gamma was constructed to express Cyprinus carpio (common carp) IFN-gamma fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-gamma with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-gamma (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 degrees C. rIFN-gamma could be recognized by western blotting with rabbit anti-grass carp IFN-gamma polydonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-gamma had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-gamma showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-gamma significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-gamma would act as an immuno-enhancer in carp aquaculture.
