Mei Peng1, Chang-Bai Dai, Yuan-Ding Chen. 1. Department of Molecular Biology, Institute of Medical Biology, Chinese Academy of Medical Sciences/Peking Union Medical College, 379 Jiaoling Road, Kunming 650118, Yunnang Province, China. mpjpm@public.km.yn.cn
Abstract
AIM: To construct and highly express an epitope of hepatitis C virus (HCV) in a foreign epitope presenting vector based on an insect virus, and to study the antigenicity of the epitope. METHODS: The HCV epitope sequence (amino acid residues 315 to 328: EGHRMAWDMMMNWS) of the E1 region was constructed at different positions of a foreign epitope presenting vector based on an insect virus, flock house virus (FHV) capsid protein encoding gene as a vector, and expressed in E. coli cells. Western blotting and ELISA were used to detect the immunoreactivity of these recombinant proteins. RESULTS: The gene encoding of the concerned B-cell epitope of HCV E1 envelope protein was expressed on FHV capsid carrier protein at positions I1 (aa 106), I2 (aa 153) and I3 (aa 305), respectively, on the surface of FHV capsid protein. The recombinant proteins in this system could be highly expressed in more than 40% of total cell protein of E. coli BL21. All the expressed recombinant proteins were in inclusion body form, and showed obvious immunoreactivity by Western blotting. Further purified recombinant proteins were detected by indirect ELISA as coating antigen respectively. All recombinant proteins could still show immunoreactivity. CONCLUSION: The epitope of HCV E1 envelope protein can be highly expressed in FHV carrier system as a chimeric protein with high immunoreactivity. This system has multiple entry sites conferring many possible conformations closer to the native one for a given sequence.
AIM: To construct and highly express an epitope of hepatitis C virus (HCV) in a foreign epitope presenting vector based on an insect virus, and to study the antigenicity of the epitope. METHODS: The HCV epitope sequence (amino acid residues 315 to 328: EGHRMAWDMMMNWS) of the E1 region was constructed at different positions of a foreign epitope presenting vector based on an insect virus, flock house virus (FHV) capsid protein encoding gene as a vector, and expressed in E. coli cells. Western blotting and ELISA were used to detect the immunoreactivity of these recombinant proteins. RESULTS: The gene encoding of the concerned B-cell epitope of HCV E1 envelope protein was expressed on FHV capsid carrier protein at positions I1 (aa 106), I2 (aa 153) and I3 (aa 305), respectively, on the surface of FHV capsid protein. The recombinant proteins in this system could be highly expressed in more than 40% of total cell protein of E. coli BL21. All the expressed recombinant proteins were in inclusion body form, and showed obvious immunoreactivity by Western blotting. Further purified recombinant proteins were detected by indirect ELISA as coating antigen respectively. All recombinant proteins could still show immunoreactivity. CONCLUSION: The epitope of HCV E1 envelope protein can be highly expressed in FHV carrier system as a chimeric protein with high immunoreactivity. This system has multiple entry sites conferring many possible conformations closer to the native one for a given sequence.
Authors: T Miyamura; I Saito; T Katayama; S Kikuchi; A Tateda; M Houghton; Q L Choo; G Kuo Journal: Proc Natl Acad Sci U S A Date: 1990-02 Impact factor: 11.205
Authors: W M Ching; C Wychowski; M J Beach; H Wang; C L Davies; M Carl; D W Bradley; H J Alter; S M Feinstone; J W Shih Journal: Proc Natl Acad Sci U S A Date: 1992-04-15 Impact factor: 11.205
Authors: G Kuo; Q L Choo; H J Alter; G L Gitnick; A G Redeker; R H Purcell; T Miyamura; J L Dienstag; M J Alter; C E Stevens Journal: Science Date: 1989-04-21 Impact factor: 47.728