葡萄球菌肠毒素C基因的克隆表达及其生物学活性鉴定

Cloning and expression of staphylococcal enterotoxin C

为表达和纯化金黄色葡萄球菌(金葡菌)超抗原毒素,本研究从金葡菌中克隆葡萄球菌肠毒素C2(sec2)基因,构建了表达葡萄球菌肠毒素C与谷胱甘肽硫转移酶(GST-SEC)融合蛋白的重组质粒(pGEMSEC),通过大肠杆菌表达、亲和纯化、酶切、再亲和纯化,建立了便于大量制备高纯度的重组SEC(rSEC)的简便方法。此外,通过酶联免疫试验及双向琼脂扩散试验检测了rSEC免疫反应性,并通过测定rSEC刺激鼠脾细胞产炎性因子来检测了rSEC的超抗原活性。结果表明,rSEC与野生型SEC具有相同的免疫反应性和超抗原活性,为在今后对该超抗原进行进一步应用性开发和构建定点减毒诱变奠定了工作基础。

In this study we cloned staphylococcal enterotoxin C2 gene （sec2） from Staphylococcus aureus and constructed a bacteria strain that expresses staphylococcal enterotoxin C and Glutathione S-transferase fusion protein. The recombinant SEC was purified and its immunoreactivity was confirmed by enzyme linked immunosorbent assay （ELISA） and double agar diffusion test. The superantigenic activity was analyzed by detecting the production of inflammatory cytokines in mouse spleen cell culture stimulated by rSEC. The results indicated that rSEC has similar biological activity with wild SEC （wSEC）.