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目的分析比较布鲁杆菌两种外膜蛋白Cu-Zn SOD和GroEL的细胞免疫特性。方法以布鲁杆菌16M基因组为模板,PCR扩增Cu-Zn SOD和GroEL蛋白的基因序列。采用Gateway方法构建表达载体,在大肠埃希菌中诱导表达并纯化,用获得的纯化蛋白刺激减毒活疫苗S19免疫小鼠的脾细胞,并测定细胞上清中IFN-γ的水平。结果成功克隆表达了Cu-Zn SOD和GroEL蛋白,用这两种蛋白刺激S19免疫小鼠脾细胞后,细胞上清中的IFN-γ水平明显升高,并且GroEL刺激后的IFN-γ水平高于Cu-Zn SOD刺激后的IFN-γ水平。结论 GroEL和Cu-Zn SOD蛋白均具有细胞免疫反应特性,且GroEL的细胞免疫反应特性强于Cu-ZnSOD。
OBJECTIVE: To analyze the cellular immunity characteristics of Brucella outer membrane proteins Cu-Zn SOD and GroEL. Methods The gene sequences of Cu-Zn SOD and GroEL were amplified by PCR using Brucella 16M genome as a template. The expression vector was constructed by Gateway method and expressed in Escherichia coli. The purified protein was used to stimulate the splenocytes of mice immunized with the attenuated live vaccine S19 and the level of IFN-γ in the supernatant was measured. Results The Cu-Zn SOD and GroEL proteins were successfully cloned and expressed. The levels of IFN-γ in the supernatants of S19-stimulated splenocytes were significantly increased after stimulated with these two proteins, and the level of IFN-γ after GroEL stimulation was high IFN- [gamma] levels after Cu-Zn SOD stimulation. Conclusion Both GroEL and Cu-Zn SOD proteins have cellular immune response characteristics, and the cellular immune response of GroEL is stronger than that of Cu-ZnSOD.