Cloning, expression, and purification of a highly immunogenic recombinant gonadotropin-releasing hormone (GnRH) chimeric peptide

Jinshu Xu, Zheng Zhu, Peng Duan, Wenjia Li, Yin Zhang, Jie Wu, Zhuoyi Hu, Rouel S. Roque, Jingjing Liu

Research output: Contribution to journalArticlepeer-review

11 Scopus citations

Abstract

To design an anti-gonadotropin-releasing hormone (GnRH) vaccine capable of eliciting strong immunogenicity, a gene fragment encoding a chimeric peptide was constructed using polymerase chain reaction and ligated into a novel expression vector for recombinant expression in a T7 RNA polymerase-based expression system. The chimeric peptide called GnRH3-hinge-MVP contained three linear repeats of GnRH (GnRH3), a fragment of the human IgG1 hinge region, and a T-cell epitope of measles virus protein (MVP). The expression plasmid contained the GnRH3-hinge-MVP construct ligated to its fusion partner (AnsB-C) via an unique acid labile Asp-Pro linker. The recombinant fusion protein was expressed in an inclusion body in Escherichia coli under IPTG or lactose induction and the target peptide was easily purified using washing of urea and ethanol precipitation. The target chimeric peptide was isolated from the fusion partner following acid hydrolysis and purified using DEAE-Sephacel chromatography. The purified GnRH3-hinge-MVP was determined to be highly homogeneous by IEF analysis and the N-terminal sequencing. Further, immunization of female mice with the recombinant chimeric peptide resulted in generation of high-titer antibodies specific for GnRH. The results showed that GnRH3-hinge-MVP could be considered as a candidate anti-GnRH vaccine.

Original languageEnglish
Pages (from-to)163-170
Number of pages8
JournalProtein Expression and Purification
Volume50
Issue number2
DOIs
StatePublished - Dec 2006
Externally publishedYes

Keywords

  • Anti-GnRH antibodies responses
  • Gonadotropin-releasing hormone (GnRH)
  • High-level expression
  • Human IgG1
  • Purification
  • T-cell epitope
  • Vaccine

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