[Expression of the human papillomavirus type 16L/E7 fusion protein in E. coli and observation of its immunogenicity in mice].
BACKGROUND
Many epidemiological and experimental evidences prove that cervical cancers are strongly associated with genital high-risk types of human papillomavirus (HPV). HPV16 is present in 50% of the tumor specimens. Thus, it is important to develop vaccines against HPV16 and cervical cancer. The authors studied the expression of the HPV16 L1DeltaCE7N fusion protein in E. coli and observed its immunogenicity.


METHODS
The fragment of HPV16 L1DeltaC gene and the E7N gene were amplified by PCR separately; the fusion gene named L1DeltaCE7N was generated by fusing E7N to the C terminal of L1DeltaC then the chimeric gene was cloned into prokaryotic expression vector pGEX-2T and expressed in E. coli strain JM109. The L1DeltaCE7N protein expressed were detected by Western blot. Finally its immunogenicity was characterized in immunized mice.


RESULTS
It was proved that the sequence and open reading frame of fusion gene L1DeltaE7N was correct by sequencing; SDA-PAGE gel analysis showed that HPV16 L1/E7 fusion protein was highly expressed in E. coli; the protein was expressed as soluble form and the molecular weight was about 85 x 10(3). The fusion protein could be purified by affinity chromatography and gel filtration. The ELISA result indicated that L1/E7 could elicit specific antibodies against L1 and E7 in immunized mice. In vivo tumor protection test indicated that tumor formation was retarded or prevented in the mice after vaccination with L1/E7, when C57 BL/6 mice were challenged by syngeneic HVP16E6 and E7 transformed tumor cells.


CONCLUSIONS
HPV16L1/E7 fusion protein was expressed in E. coli, it can be a candidate for prophylactic and therapeutic vaccine for HPV16-associated infection and tumors.
Tian
Hou-wen
HW
Institute for Viral Disease Control and Prevention, China CDC, Beijing 100052, China. houwentian@yahoo.com.cn
Ye
Zhen-mei
ZM
Lu
Zhen-hua
ZH
Jao
Ren
R
Bian
Tao
T
Zhao
Li
L
Ruan
Li
L
D000818:Animals; D015153:Blotting, Western; D045744:Cell Line, Tumor; D004926:Escherichia coli / Q000235:genetics*; D005260:Female; D006801:Humans; D007114:Immunization / Q000379:methods; D051379:Mice; D008810:Mice, Inbred C57BL; D009368:Neoplasm Transplantation; D009374:Neoplasms, Experimental / Q000276:immunology / Q000473:pathology / Q000517:prevention & control; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D027383:Papillomaviridae / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D030361:Papillomavirus Infections / Q000276:immunology / Q000473:pathology / Q000517:prevention & control; D053918:Papillomavirus Vaccines / Q000008:administration & dosage / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology* / Q000378:metabolism / Q000648:ultrastructure
C424202:HPV16 L1E7 chimeric virus-like particle; D015514:Oncogene Proteins, Fusion; D009856:Oncogene Proteins, Viral; D053918:Papillomavirus Vaccines; D011993:Recombinant Fusion Proteins