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目的确定肺炎链球菌自溶酶(LytA)蛋白B细胞线性表位。方法通过生物信息学方法预测肺炎链球菌LytA蛋白分子的B细胞表位并人工合成相应肽段;经克隆、表达、纯化等步骤得到纯化的重组LytA(rLytA)蛋白用以免疫小鼠并进行Westen blot鉴定;取免疫组小鼠和对照组小鼠血清分别与人工合成的候选B细胞表位肽段进行间接ELISA检测,筛选出B细胞表位。结果利用多种方法分析LytA蛋白分子的B细胞表位,经整理得到11条候选肽段,命名为LB1到LB11,并进行了人工合成。利用分子生物学技术获得了rLytA蛋白并成功地免疫了小鼠,间接ELISA结果显示:肽段LB3、LB5、LB6、LB7、LB11与rLytA免疫小鼠的血清发生反应,其OD 450 nm读数比相应的阴性对照小鼠显著升高,差异有统计学意义(P<0.01)。结论确定了肺炎链球菌LytA蛋白分子5个B细胞线性表位的位置和序列。
Objective To determine the linear epitopes of Streptococcus pneumoniae autolysin (LytA) protein B cells. Methods Bioinformatics methods were used to predict the B cell epitopes of LytA protein of S. pneumoniae and to synthesize the corresponding peptides. The purified recombinant LytA (rLytA) protein was cloned, expressed and purified to immunize mice and Westen blot identification; take the immune mice and control mice serum respectively with synthetic candidate B cell epitope peptide indirect ELISA test to screen B cell epitopes. Results A number of methods were used to analyze the B cell epitopes of LytA protein molecules. Eleven candidate peptides were obtained and named as LB1 to LB11 and synthesized. The rLytA protein was obtained by molecular biology technique and the mice were successfully immunized. The indirect ELISA results showed that the peptides of LB3, LB5, LB6, LB7 and LB11 reacted with the serum of mice immunized with rLytA, Negative control mice significantly increased, the difference was statistically significant (P <0.01). Conclusion The location and sequence of five B-cell linear epitopes of Streptococcus pneumoniae LytA protein molecule were determined.