| 胡美丽,刘洪洋,王志荣,陈 雪,张 婷,许雪梅.原核表达系统制备HPV58L1 VLP可诱发高滴度且持久的中和抗体[J].,2018,(3):419-422 |
| 原核表达系统制备HPV58L1 VLP可诱发高滴度且持久的中和抗体 |
| Human Papillomavirus Type 58 L1 Virus-Like Particles Produced in Prokaryotic System Elicit High Level of Long-lasting Specific Neutralizing Antibodies |
| 投稿时间：2017-05-23 修订日期：2017-06-18 |
| DOI：10.13241/j.cnki.pmb.2018.03.004 |
| 中文关键词: 人乳头瘤病毒58型 大肠杆菌表达系统 病毒样颗粒 中和抗体 |
| 英文关键词: Human papillomavirus type 58 Escherichia coli expression system Virus-like particles Neutralizing antibody |
| 基金项目:国家国际科技合作专项（2013DFA32430）；中国医学科学院医学与健康科技创新工程基金项目（2016-I2M-3-026） |
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| 中文摘要: |
| 摘要 目的：采用大肠杆菌表达系统制备人乳头瘤病毒58型(human papillomavirus type 58，HPV58)病毒样颗粒(virus-like particle，VLP)疫苗。方法：合成法获得HPV58 L1大肠杆菌密码子优化基因，构建HPV58 L1重组原核表达质粒mpET22b/HPV58 L1，检测其在BL21(DE3)中表达水平，饱和硫酸铵沉淀加阳离子交换层析法纯化蛋白后进行动态光散射(dynamic light scatter，DLS)分析。小鼠免疫后，检测免疫血清针对HPV58假病毒的中和抗体水平。结果：HPV58 L1蛋白在BL21(DE3)细胞中大部分以可溶形式表达，纯化获得的HPV58 L1蛋白可组装成水动力学直径约为74 nm的VLP。0.5 μg的HPV58 L1 VLP可诱发小鼠产生高滴度的HPV58特异性中和抗体，可维持至少20周。结论：原核表达系统制备的HPV58 L1 VLP可诱发高滴度且持久的中和抗体，可用于成本低的HPV58疫苗的研究。 |
| 英文摘要: |
| ABSTRACT Objective: To produce human papillomavirus type 58 (HPV58) virus-like particles (VLP) vaccine with Escherichia coli expression system. Methods: A synthesized HPV58 L1 gene optimized with bias condons of Escherichia coli was inserted into prokaryotic expression vector mpET22b, and expressed in BL21(DE3). Cells were sonicated and the supernatent was purified with ammonium sulfate precipitation plus cation-exchange chromatography. The purified HPV58 L1 protein was analyzed by dynamic light scatter (DLS), and then immunized mice twice with two week interval. Sera were analyzed with pseudovirion-based neutralization assays. Results: The HPV58 L1 protein was expressed largely in cell lysate and was easy to purify. Dynamic light scattering analysis showed that the mean hy- drodynamic diameter of the purified protein was 74 nm, which was similar to that of HPV58 L1 VLP control, indicating the protein was self-assembled into HPV58 L1 VLP. The mean titer of HPV58 neutralizing antibody was as high as 10,000 in sera collected at week 4, and retained 5,000 at week 22. Conclusion: The HPV58 Ll VLP produced in prokaryotic system can induce high titer of long-lerm spe- cific neutralizing antibody, which is a cost-effective candidate of HPV58 prophylatic vaccine. |
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