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目的预测和鉴定梅毒螺旋体(Tp)膜蛋白TprF氨基端保守区(TprFN)的优势B细胞表位,为深入研究梅毒多价表位疫苗提供依据。方法从GenBank获取TprFN的氨基酸序列,采用Mobyle、ABCpred和IEDB在线软件综合分析预测TprFN的B细胞表位并人工合成多肽;表达重组蛋白TprFN并经Western blot鉴定后免疫兔,获取血清并测定抗体效价;以TprFN免疫兔血清、梅毒患者血清(设正常人血清和正常兔血清为阴性对照),间接ELISA测定预测的7条人工合成的B细胞表位多肽的免疫反应性和特异性。结果软件综合预测TprFN的P1(43-62AA)、P2(57-71AA)、P3(81-88AA)、P4(89-103AA)、P5(125-138AA)、P6(231-251AA)、P7(268-279AA)可能为B细胞表位;表达一可溶性蛋白,WB鉴定为目的蛋白,其免疫抗体效价为1∶12 800以上;ELISA结果显示,预测表位P1、P3与TprFN免疫兔血清及梅毒患者血清均呈阳性反应,而与对照血清均不反应。结论 P1、P3为TprF潜在的优势B细胞表位。
Objective To predict and identify the dominant B cell epitopes of the TprF conserved region (TprFN) of membrane protein TprF, and to provide basis for further study of multivalent epitope vaccine of syphilis. Methods The amino acid sequence of TprFN was obtained from GenBank. The software was used to predict the B cell epitope of TprFN by using the software of Mobyle, ABCpred and IEDB. The recombinant protein TprFN was expressed and identified by Western blot. The serum was obtained and the antibody titer was determined The immunoreactivity and specificity of the predicted seven synthetic B-cell epitope polypeptides were determined by indirect ELISA using the serum of rabbits immunized with TprFN and the serum of syphilis patients (normal human serum and normal rabbit serum as negative control). Results The software was able to predict P1 (43-62AA), P2 (57-71AA), P3 (81-88AA), P4 (89-103AA), P5 (125-138AA), P6 268-279AA) may be B cell epitopes; expressed a soluble protein, WB identified as the target protein, the antibody titer of 1:12 800 or more; ELISA results show that predicting the epitope P1, P3 and TprFN immune rabbit serum and Serum syphilis patients were positive, but not with the control serum. Conclusion P1 and P3 are the potential predominant B cell epitopes of TprF.