title: Protective immunity of Pichia pastoris-expressed recombinant envelope protein of Japanese encephalitis virus.
abstract: Japanese encephalitis virus (JEV) envelope (E) protein holds great promise for use in the development of a recombinant vaccine. Purified recombinant E (rE) protein may be useful for numerous clinical applications; however, there are limitations in using the Escherichia coli expression system for producing high-quality rE protein. Therefore, in this study, the yeast expression system was used to generate the rE protein. For protein production using the yeast system, the full-length JEV E gene was cloned into Pichia pastoris. SDS-PAGE and immunoblotting analysis demonstrated that the rE protein had a molecular mass of 58 kDa and was glycosylated. The predicted size of the mature unmodified E protein is 53 kDa, suggesting that post-translational modifications resulted in the higher molecular mass. The rE protein was purified to greater than 95% purity using combined ammonium sulfate precipitation and a SP-Sepharose Fast Flow column. This purified rE protein was evaluated for immunogenicity and protective efficacy in mice. The survival rates of mice immunized with the rE protein were significantly increased over that of Hyphantria cunea nuclear polyhedrosis virus E protein (HcE). Our results indicate that the rE protein expressed in the P. pastoris expression system holds great promise for use in the development of a subunit vaccine against JEV.
authors.affiliation: Department of Environmental Health and Safety, Eulji University, Sungnam 461-713, Korea.
authors.forename: Woo-Taeg
authors.initials: WT
authors.lastname: Kwon
authors.forename: Woo-Sik
authors.initials: WS
authors.lastname: Lee
authors.forename: Pyo-Jam
authors.initials: PJ
authors.lastname: Park
authors.forename: Tae-Kyu
authors.initials: TK
authors.lastname: Park
authors.forename: Hyun
authors.initials: H
authors.lastname: Kang
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology; D004664:Encephalitis Virus, Japanese / Q000235:genetics / Q000276:immunology; D004672:Encephalitis, Japanese / Q000276:immunology* / Q000401:mortality / Q000517:prevention & control* / Q000821:virology; D015870:Gene Expression; D007114:Immunization; D051379:Mice; D010843:Pichia / Q000235:genetics* / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification
chemical_list: D000914:Antibodies, Viral; D014759:Viral Envelope Proteins