HPV 6b结构蛋白L1和沙眼衣原体CTL表位嵌合病毒样颗粒在昆虫细胞中的表达

目的为研制能同时防治人乳头瘤病毒(HPV)和沙眼衣原体(Ct)的嵌合疫苗。方法将Ct主要外膜蛋白(MOMP)插入到HPV衣壳蛋白L1羧基端,通过PCR、酶切、连接等方法构建了杆状病毒表达载体,进一步在大肠杆菌中组装成重组杆状病毒,经感染昆虫细胞表达嵌合病毒样颗粒,并经SDS-PAGE电泳和Westernblot分析鉴定,电镜负染。结果嵌合蛋白HPV6bL1/CtMOMP249-268在昆虫细胞中得到了表达,表达产物的相对分子质量为56000,与HPV6bL1单抗产生特异性反应,电子显微镜观察到病毒样颗粒形成。结论在昆虫细胞中表达的含HPV6bL1和CtCTL表位的嵌合蛋白,能自行组装成病毒样颗粒,为人乳头瘤病毒和沙眼衣原体嵌合疫苗的研究奠定基础。

Construction and expression of chimeric protein of human papillomavirus type 6b L1 and CTL epitope of MOMP of Chlamydia trachomatis

To obtain the vaccine against infection of human papillomavirus and Chlamydia trachomatis,the recombinant vaccine was designed by inserting CTL epitopes of the Chlamydia trachomatis major outer membrane protein into carboxyl-terminal of HPV type 6b L1,due to the cause of HPV L1 assembling into virus like particles will not affect when the external gene was inserted into carboxyl-terminal of HPV L1.A baculovirus transfer vector including the aim gene was constructed by PCR amplifying,restriction endonuclease digestion and ligation.The recombinant baculovirus was generated in E.coli.The chimeric virus like particles was obtained by the infection and expression of recombinant baculovirus in sf-9 insect cell,analysed by SDS-PAGE and Western blot and observed by electron microscope.It indicated that the chimeric protein HPV 6b L1/Ct MOMP could be expressed in sf-9 insect cells,SDS-PAGE showed that the relative molecular weight of this Chimeric protein was about 56 KDa,and antigenic specificity was confirmed by Western blot.Electron microscope showed that the chimeric protein could assembled into virus like particles.As a conclusion,the chimeric protein including human papillomavirus type 6b L1 and CTL epitope of Chlamydia trachomatis can be efficiently expressed in sf-9 insect cells with baculovirus express system,and it can assemble itself into virus like particles.which will be valuable for the research of chimeric vaccine against the infection of with human papillomavirus and chlamydia trachomatis.