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肺炎球菌溶血素(PL)是一种巯基激活的细胞溶解毒素,它可由各型肺炎球菌产生。本研究旨在使用重组PL(rPL)作为18C型肺炎球菌多糖抗原(PS18C)的蛋白载体,建立一种T细胞依赖性、免疫原性较强的新型肺炎球菌菌苗。 将编码PL的2.8kb BstYI基因插入质粒pKK232-8中,然后转化大肠杆菌,rPL以谷胱甘肽-S转移酶融合蛋白形式在大肠杆菌中表达,通过亲和层析和羟磷灰石层析法纯化rPL。对纯化rPL分子量、等电点(pI)、氨基酸顺序以及生物学和免疫学特性进行了研究。此外,作者通过还原氨化法或间隔分子己二酸二酰肼(ADH)将纯化的rPL与PS18C结合制成两种偶联菌苗PS18C-rPL和PS18C(AH)-rPL,以小鼠为动物模型探讨这两种偶联菌苗的免疫原性及免疫保护性。
Pneumolysin (PL) is a sulfhydryl-activated cytolytic toxin that can be produced by various types of pneumococci. This study aimed to establish a T-cell dependent and immunogenic pneumococcal vaccine using recombinant PL (rPL) as a protein carrier for the 18C pneumococcal polysaccharide antigen (PS18C). The 2.8 kb BstYI gene encoding PL was inserted into plasmid pKK232-8 and then transformed into E. coli. The rPL was expressed in E. coli as a glutathione-S-transferase fusion protein and purified by affinity chromatography and hydroxyapatite layer Analytical purification of rPL. Purified rPL molecular weight, isoelectric point (pI), amino acid sequence and biological and immunological properties were studied. In addition, the authors combined the purified rPL with PS18C by reductive amination or the spacer molecule adipic acid dihydrazide (ADH) to make two coupled bacterins, PS18C-rPL and PS18C (AH) -rPL, in mice Animal models to explore the immunogenicity and immunoprotection of these two coupled vaccine.