title: Secretory Expression of Different C-terminal Truncated HCV E1 Proteins in Mammalian Cells and Characterization of the Expressed Products.
abstract: Three fragments of the HCV envelope 1 (E1) with different C-terminal truncation at aa310, aa325, aa340 were cloned into the mammalian expression vector pSecTagB. An epitope in the hepatitis B surface antigen, preS1(21--47), were genetically engineered onto the N-terminus of the recombinant protein and used as an affinity tag for detection and purification. The resulting pSec-preS1-E1t310, pSec-preS1-E1t325 and pSec-preS1-E1t340 were transiently expressed in the HeLa cells and the antigenicity, secretory efficiency and glycosylation type of the recombinant E1 proteins were compared. All of the three recombinant proteins could be detected by both preS1 monoclonal antibody and E1 polyclonal antiserum. The expression products were secreted and highly mannose-type glycosylated, with S1E1t325 being secreted, indicating the influence of the hydrophobic regions on the secretion of the E1 protein. Three CHO cell lines expressing the proteins, S1E1t310, S1E1t325 and S1E1t340, were established and the CHO/pSecS1E1t325 was chosen for further study. The secreted S1E1t325 could be enriched from cell culture medium by the preS1 antibody-coupled Sepharose. The glycosylation analysis indicated the lack of complex glycogen even after the E1 was secreted via Golgi complexes. The established stable cell lines and anti-preS1 affinity method could be utilized to enrich and purify the HCV E1 expressed in mammalian cells, and may be used for further characterization of this protein.
authors.affiliation: Institute of Biochemistry and Cell Biology, Shanghai Institute for Biological Sciences, the Chinese Academy of Sciences, Shanghai 200031, China. wangyuan@server.shcnc.ac.cn
authors.forename: J
authors.initials: J
authors.lastname: Zhu
authors.forename: Y Y
authors.initials: YY
authors.lastname: Kong
authors.forename: J
authors.initials: J
authors.lastname: Liu
authors.forename: Z C
authors.initials: ZC
authors.lastname: Zhang
authors.forename: Y
authors.initials: Y
authors.lastname: Wang
authors.forename: G D
authors.initials: GD
authors.lastname: Li