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Improving antigenicity of the recombinant hepatitis C virus core protein via random mutagenesis.

ABSTRACT: In order to enhance the sensitivity of diagnosis, a recombinant clone containing domain I of HCV core (amino acid residues 1 to 123) was subjected to random mutagenesis. Five mutants with higher sensitivity were obtained by colony screening of 616 mutants using reverse ELISA. Sequence analysis of these mutants revealed alterations focusing on W(84), P(95), P(110), or V(129). The inclusion bodies of these recombinant proteins overexpressed in E. coli BL21(DE3) were subsequently dissolved using 6 M urea and then refolded by stepwise dialysis. Compared to the unfolded wild-type antigen, the refolded M3b antigen (W(84)S, P(110)S and V(129)L) exhibited an increase of 66% antigenicity with binding capacity of 0.96 and affinity of 113??M(-1). Moreover, the 33% decrease of the production demand suggests that M3b is a potential substitute for anti-HCV antibody detection.

SUBMITTER: Huang CJ

PROVIDER: S-EPMC3166572 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Improving antigenicity of the recombinant hepatitis C virus core protein via random mutagenesis.

Huang Chen-Ji CJ Peng Hwei-Ling HL Cheng Chih-Yu CY

Journal of biomedicine & biotechnology 20110829

In order to enhance the sensitivity of diagnosis, a recombinant clone containing domain I of HCV core (amino acid residues 1 to 123) was subjected to random mutagenesis. Five mutants with higher sensitivity were obtained by colony screening of 616 mutants using reverse ELISA. Sequence analysis of these mutants revealed alterations focusing on W(84), P(95), P(110), or V(129). The inclusion bodies of these recombinant proteins overexpressed in E. coli BL21(DE3) were subsequently dissolved using 6 ...[more]

PMID: 21904443

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