Expression, purification and immunogenicity analysis of HPV type 18 virus-like particles from Escherichia coli.
- Author:
Minghui XIE
1
;
Shaowei LI
;
Wentong SHEN
;
Zhongyi LI
;
Yudi ZHUANG
;
Xiaobing MO
;
Ying GU
;
Ting WU
;
Jun ZHANG
;
Ningshao XIA
Author Information
1. National Institute of Diagnostics and Vaccine Development in Infectious Diseases, Key Laboratory of the Ministry of Education for Cell Biology and Tumor Cell Engineering, School of Life Science, Xiamen University, Xiamen 361005, China.
- Publication Type:Journal Article
- MeSH:
Animals;
Capsid Proteins;
biosynthesis;
genetics;
immunology;
Escherichia coli;
genetics;
metabolism;
Goats;
Human papillomavirus 18;
immunology;
isolation & purification;
Mice;
Oncogene Proteins, Viral;
biosynthesis;
genetics;
immunology;
Rabbits;
Recombinant Proteins;
biosynthesis;
genetics;
immunology;
Virion;
genetics;
immunology
- From:
Chinese Journal of Biotechnology
2009;25(7):1082-1087
- CountryChina
- Language:Chinese
-
Abstract:
Here, we presented a method to bacterially express the major structural protein L1 of Human Papillomavirus type 18 (HPV18) as soluble form. We found that the purified L1 could self-assemble to virus-like particles (VLPs). Further, we investigated the immunogenicity and the induced level of neutralizing antibody using these VLPs. First, the genome of HPV18 was cloned from a patient in Xiamen. It was used as template for PCR amplification of HPV18 L1 gene. The resultant DNA fragment was inserted into expression vector pTrxFus and expressed in Escherichia coli GI724. Second, L1 protein was purified by ammonium sulfate precipitation, ion-exchange chromatography and hydrophobic interaction chromatography; and the purified L1 was subjected to self-assembly to form VLPs with the removal of premixed reductant DTT. Finally, the size and morphology of these VLPs was investigated by Dynamic Light Scattering and Transmission Electronic Microscopy as 29.34 nm in hydrated radius and globular particles similar with native HPV18. The half effective dosage (ED50) and maximum level of neutralizing antibody elicitation were measured by vaccinations on mice, rabbit and goat using pseudovirus neutralization cell model. The results showed that the ED50 of HPV18 VLPs is 0.006 microg in mice, and the maximum titer of neutralizing antibody elicited in rabbit and goat is up to 10(7). As a conclusion, we can provide HPV18 VLPs with highly immunogenicity from prokaryote expression system, which may pave a new way for research and development of prophylactic vaccine for HPV18.
