中国明对虾(Fenneropenaeus chinensis)丝氨酸蛋白酶同源物基因(Fc-SPH)的重组表达及活性分析

RECOMBINANT EXPRESSION AND BIOACTIVITY ANALYSIS OF A SERINE PROTEINASE HOMOLOGUE GENE (Fc-SPH) FROM THE CHINESE SHRIMP FENNEROPENAEUS CHINENSIS

以实验室前期克隆到的中国明对虾丝氨酸蛋白酶同源物基因(Fc-SPH)(GenBank登录号:DQ318859)为基础,利用原核表达系统对Fc-SPH基因成熟肽区域进行了重组表达和纯化复性分析,并对获得的重组目的蛋白开展了抑菌活性及微生物清除功能研究。结果表明:在体外成功获得了大量有活性的对虾Fc-SPH蛋白(rFc-SPH),活性研究显示rFc-SPH对大多数受试菌种都有明显的抑制效果,并可以加速中国明对虾清除体内外源微生物的速度。作者推断Fc-SPH即可以作为一线防御应答效应物直接参与虾类的先天免疫活动而发挥作用,也可以通过调理作用促进血细胞对病原微生物的吞噬和杀灭。

Like other crustaceans, shrimp have a non-specific innate immunity which is composed of diverse processes and molecules to defend themselves against invading pathogens. Arthropod clip domain serine proteinase homologues （clip-SPHs） have been shown to be involved in various biological functions including innate immunity. A serine proteinase homologue （Fc-SPH） cDNA was cloned from the haemocytes ofFenneropenaeus chinensis. Fc-SPH contains a trypsin-like serine protease domain （Tryp_SPc domain） and a clip domain. The transcript of Fc-SPH is induced in response to white spot syndrome virus （WSSV） infection. Recombinant Fc-SPH （rFc-SPH） were successfully expressed in bacteria and puri- fied for further research of biological functions. It showed in vitro antimicrobial activity against the Bacillus thurigiensis, Bacillus subtilis, Vibrio anguillarum, Escherichia coli, l/ibrio harveyi and Saccharomyces cerevisiae. A higher bacterial clearance rate of E harveyi coated with the rFc-SPH compared with E harveyi only was shown by in vivo study. It revealed that Fc-SPH act as opsonin and could help shrimp to remove external microorganism. The results suggest that Fc-SPH might be a multifunctional immune molecule in shrimp defense. It can provide us clues to understand the probable role of SPH in innate immunity of shrimp.