斯氏并殖吸虫蛋白质组学研究

发布时间：2018-09-19 11:23

【摘要】：目的对斯氏并殖吸虫(Paragonimus skrjabini)囊蚴、童虫和成虫蛋白质组进行研究。方法在贵州省斯氏并殖吸虫疫源地开阳、毕节采集溪蟹,从中分离囊蚴置于PBS中,灌胃感染SD大鼠3只(10个囊蚴/只),喂食感染雄性犬3只(20个囊蚴/只)。感染大鼠后1个月解剖取童虫,感染犬后3个月解剖取成虫。将囊蚴、童虫和成虫中加入中性组织裂解液,冰上超声破碎提取总蛋白,Bradford法测定蛋白含量并进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)及双向凝胶电泳,采用PDquest 8.0软件对双向凝胶电泳图像进行分析寻找差异点。对差异点进行酶解、质谱鉴定。最后进行NCBI在线及本地数据库检索。结果 SDS-PAGE显示囊蚴、童虫和成虫总蛋白主要集中分布在相对分子质量(Mr)25 000~116 000。双向凝胶电泳共筛选出51个差异蛋白点,分别为囊蚴20个、童虫25个、成虫6个。因成虫差异点灰度值两两间的比值变化远小于囊蚴与童虫,故未进行质谱鉴定。36个囊蚴和童虫的肽段序列进行分析后发现,在线检索蛋白种类主要为无色杆菌(Achromobacter lyticus)蛋白酶Ⅰ(一种赖氨酸特异性的丝氨酸酶)、脱氢抗坏血酸还原酶蛋白、谷胱甘肽S转移酶DHAR2类似物、热激蛋白Hsp82及Hsp96-β、肌动蛋白和半胱氨酸酶抑制剂等;本地复殖目数据库检索主要蛋白种类是半胱氨酸酶、肌动蛋白和热激蛋白。结论斯氏并殖吸虫囊蚴与童虫蛋白组成有差异,囊蚴含有肌动蛋白,童虫含有解毒及应激蛋白,两者都含有水解酶类及半胱氨酸酶。
[Abstract]:Objective to study the proteome of paragonimus skrjabini (Paragonimus skrjabini) cysticercus, juvenile worm and adult worm. Methods in Kaiyang, Guizhou Province, where paragonimus skrjabini foci were collected, the cysticercus was isolated and placed in PBS. Three SD rats (10 cysticercaria / cysticercaria) were fed with infected male dogs (20 cysticercaria / rat). The adult worms were dissected 1 month after infection and 3 months after infection. The neutral tissue lysate was added to cysticercus, juvenile worm and adult. The total protein was extracted by ultrasonic crushing on ice. The protein content was determined by SDS-PAGE and two dimensional gel electrophoresis. The two-dimensional gel electrophoresis image was analyzed by PDquest 8.0 software to find out the difference point. The differential sites were hydrolyzed by enzyme and identified by mass spectrometry. Finally, NCBI online and local database retrieval. Results SDS-PAGE showed that the total proteins of cysticercus, juvenile worm and adult were mainly distributed in the relative molecular weight of (Mr) 25 000 ~ 116 000. A total of 51 differential protein spots were screened by two dimensional gel electrophoresis, including 20 cysticercaria, 25 juvenile worms and 6 adult worms. Because the ratio of gray value of the difference point of adult is much smaller than that of cysticercus and child worm, it is not identified by mass spectrometry. After analyzing the peptide sequence of 36 cysticercus and child worm, it was found that, The online search proteins are mainly (Achromobacter lyticus) protease I (a lysine-specific serine enzyme), dehydroascorbic acid reductase protein and glutathione S-transferase DHAR2 analogue. Heat shock protein (Hsp82) and Hsp96- 尾, actin and cysteinase inhibitor were found in the local colonization database, and the main proteins were cysteinase, actin and heat shock protein. Conclusion there are differences in protein composition between metacercaria of paragonimus skrjabini and that of child worm. Actinomycin is found in cysticercaria and detoxification and stress protein in juvenile worm. Both of them contain hydrolases and cysteinase.
【作者单位】：贵州省疾病预防控制中心;贵州师范大学生命科学院;
【基金】：贵州省优秀科技教育人才省长资金项目(No.2010-91) 贵州省科学技术基金(No.[2012]2210)~~
【分类号】：R383.23

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