肺炎嗜衣原体Cpn0797蛋白的生物信息学分析及其单克隆抗体制备鉴定

Bioinformatic analysis of Chlamydophila pneumoniae Cpn0797 protein and the preparation of its monoclonal antibody

目的生物信息学分析肺炎嗜衣原体Cpn0797蛋白的结构,制备特异性的抗Cpn0797蛋白的单克隆抗体。方法用ProtParam、SignalP、NPS@、和PSORT等软件对Cpn0797蛋白的理化参数、信号肽、二级结构、蛋白亚细胞定位进行分析;原核表达纯化GST-Cpn0797融合蛋白,以其为免疫原免疫BALB/c小鼠,杂交瘤技术制备单克隆抗体,采用间接免疫荧光法鉴定单克隆抗体的亚类和特异性。结果生物信息学分析表明Cpn0797蛋白二级结构以无规则卷曲为主;成功地建立了能稳定分泌抗Cpn0797蛋白的单克隆抗体杂交瘤细胞株,单克隆抗体能特异性的识别Cpn0797内源性蛋白。结论成功制备特异性的Cpn0797单克隆抗体,为进一步探究Cpn0797蛋白的生物学功能提供实验基础。

In order to analyze the protein structure of Chlamydophila pneurnoniae Cpn0797 protein and to prepare spe cific monoclonaI antibodies （mAb） against Cpn0797 protein, ProtParam, SignalP, NPS@ and PSORT software packages were used to predict the physical and chemical properties, signal peptide, secondary structure and protein localization sites in cells according to the amino acid sequence of Cpn0797. The gene cpn0797 was cloned into the prokaryotic expression vector pGEX6p-2, and expressed in E. coli and then purified. The mAbs against Cpn0797 were prepared with the hybridoma technique after mice were immunized with purified GSTCpn0797 fusion protein. The isotype and specificities of the mAbs were determined by indirect immunofluorescence assay （IFA）. The bioinformatical analysis results showed that Cpn0797 protein mainly consisted of random coils. Hybridoma cell lines stably secreting mAbs against Cpn0797 protein are obtained. The mAbs reacted with Cpn0797 endogenous protein. In conclusion, the specific mAbs against Cpn0797 protein were obtained, which pro- vides a basis for further study in the biological function of Cpn0797 protein.