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目的预测和鉴定梅毒螺旋体(Treponema pallidum,Tp)Tp92蛋白的Th细胞表位,为深入探讨这些表位在梅毒表位疫苗中的作用奠定基础。方法采用RANKPEP和SYFPEITHI软件分析工具联合预测Tp92蛋白的Th细胞表位,人工合成5条表位多肽,以5条表位多肽(同时设Con A阳性对照和RPMI-1640阴性对照)分别刺激梅毒患者和正常人外周血单个核细胞(PBMC),用CCK-8法鉴定预测的Th细胞表位,进一步以ELISA检测INF-γ和IL-4的分泌水平,区分Th细胞表位的类型。结果软件预测显示,Tp92蛋白的P1(103-118AA)、P2(694-712AA)、P3(668-680AA)、P4(300-313AA)、P5(396-410AA)氨基酸序列最可能为其Th细胞表位。CCK-8法检测显示,P2、P3、P5能诱导梅毒患者淋巴细胞明显增殖,而正常人淋巴细胞不反应;ELISA检测细胞因子产生情况发现:P2、P3、P5可刺激梅毒患者淋巴细胞产生较高水平IFN-γ,而不产生IL-4。结论 P2、P3、P5为Tp92蛋白潜在的HLA-DRBl限制性特异性Th1型细胞表位。
Objective To predict and identify the Th cell epitopes of Tp92 protein of Treponema pallidum (Tp), and lay a foundation for further exploration of the role of these epitopes in the syphilis epitope vaccine. Methods The Th epitopes of Tp92 protein were predicted by RANKPEP and SYFPEITHI software tools. Five epitope peptides were synthesized and 5 epitopes (Con A positive control and RPMI-1640 negative control) were used to stimulate syphilis patients And peripheral blood mononuclear cells (PBMCs) from healthy volunteers. The predicted Th cell epitopes were identified by CCK-8 assay, and the levels of INF-γ and IL-4 secreted by ELISA were detected by ELISA. Results The software predicts that the amino acid sequences of P1 (103-118AA), P2 (694-712AA), P3 (668-680AA), P4 (300-313AA), and P5 (396-410AA) of Tp92 protein are most likely to be Th cells gauge. CCK-8 assay showed that P2, P3 and P5 could induce lymphocyte proliferation in syphilis patients, while normal lymphocytes did not respond. ELISA to detect the production of cytokines showed that P2, P3 and P5 stimulated the production of lymphocytes in patients with syphilis High levels of IFN-γ, but not IL-4. Conclusion P2, P3 and P5 are the potential HLA-DRB1-restricted Th1-type epitopes of Tp92 protein.