原核高效制备人乳头瘤病毒16型L1病毒样颗粒

目的通过原核表达系统高效制备人乳头瘤病毒(HPV)16晚期蛋白L1病毒样颗粒(VLPs)。方法构建HPV16L1基因序列优化前后的PET30aHPV16L1重组质粒,转化大肠杆菌Rosetta(DE3);用IPTG诱导目的基因表达,两步层析方法纯化HPV16L1蛋白;电镜下观察纯化产物形成VLPs的情况。结果成功构建大肠杆菌工程菌,高效可溶表达(目的蛋白约占总蛋白的38%)并纯化HPV16L1蛋白,纯度达95%以上,电镜下观察,发现纯化后的目的蛋白为直径50 nm左右,形态与天然病毒颗粒高度相似。结论在大肠杆菌原核系统中高效、简易地制备了HPV16L1VLPs,为诊断试剂和疫苗的研制奠定基础。

Efficient preparation of human HPV16L1 virus-like particles in prokaryotic system

Objective To efficiently prepare HPV16L 1 virus-like particles （VLPs） in Escherichia coli expression system. Methods Recombinant plasmids PET30a-HPV16L1 with condon usage optimization and non-optimization were constructed, and then transformed into E.coli Rosetta （DE3）. IPTG was used to induce the recombinant E.coli Rosetta （DE3）. By two step chromatography, the protein was purified. Electron microscopy was used to observe the structure of VLPs after purification. Results The recombinant engineered bacteria were successfully constructed. The solvable HPV16L1 protein （about 38% of the total protein） were efficiently expressed and purified, which reached the purity of higher than 95%. Self-assembling of VLPs for the purified L1 protein with the diameter of 50 nm was observed, showing high similarity with natural VLPs. Conclusion HPV16L1 VLPs was efficiently and conveniently prepared in Escherichia coli expression system, which will lay a foundation for further study of diagnostic reagent and vaccine.