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Toxoplasma gondii: expression and characterization of a recombinant protein containing SAG1 and GRA2 in Pichia pastoris

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Abstract

Toxoplasma gondii is an obligate intracellular protozoan which infects most species of warm-blooded animals and causes toxoplasmosis. Previous immunological and immunization studies have demonstrated the potential role of T. gondii antigens SAG1 and GRA2 as a vaccine candidate. In the present study, we have cloned, expressed, and purified a recombinant protein SAG1–GRA2 in Pichia pastoris. Results showed that P. pastoris was a robust system producing a large amount of highly purified and biological activity protein. BALB/c mice immunized with SAG1–GRA2 elicited stronger humoral and cellular responses in comparison to control groups. This immunization resulted in an enhanced Th1 immune response as measured by IgG2a antibody production and increased splenocyte IFN-γ production, whereas no IL-4 was detected. After a lethal challenge with the highly virulent T. gondii RH strain, a prolonged survival time in SAG1–GRA2-immunized mice was observed in comparison to control groups. Our data demonstrate that SAG1–GRA2 triggered a protective response against toxoplasmosis. Therefore, SAG1–GRA2 protein might be a good candidate for the further development of a multiantigenic vaccine.

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References

  • Alexandre J, Jebbari H, Bluethmann H, Satoskar A, Roberts CW (1996) Immunological control of T. gondii and appropriate vaccine design. Curr Top Microbiol Immunol 219:188–190

    Google Scholar 

  • Bhopale GM (2003) Development of a vaccine for toxoplasmosis: current status. Microbes Infect 5:457–462

    Article  PubMed  CAS  Google Scholar 

  • Bout DT, Mevelec MN, Velge-Roussel F, Dimier-Poisson I, Lebrun M (2002) Prospects for a human Toxoplasma vaccine. Curr Drug Targets Immune Endocr Metabol Disord 2:227–234

    Article  PubMed  CAS  Google Scholar 

  • Brinkmann V, Remington JS, Sharma SD (1993) Vaccination of mice with the protective F3G3 antigen of Toxoplasma gondii activates CD4+ but not CD8+ cells and induces Toxoplasma specific IgG antibody. Mol Immunol 30:353–358

    Article  PubMed  CAS  Google Scholar 

  • Bulow R, Boothroyd JC (1991) Protection of mice from fatal Toxoplasma gondii infection by immunization with p30 antigen in liposomes. J Immunol 147:3496–3500

    PubMed  CAS  Google Scholar 

  • Burg JL, Perelman D, Kasper LH, Ware PL, Boothroyd JC (1988) Molecular analysis of the gene encoding the major surface antigen of Toxoplasma gondii. J Immunol 141:3584–3591

    PubMed  CAS  Google Scholar 

  • Buxton D, Innes EA (1995) A commercial vaccine for ovine toxoplasmosis. Parasitology 110:S11–S16

    Article  PubMed  Google Scholar 

  • Buxton D, ThomsonK, Maley S, Wright S, Bos HJ (1991) Vaccination of sheep with a live incomplete strain (S48) of Toxoplasma gondii and their immunity to challenge when pregnant. Vet Rec 129:89–93

    PubMed  CAS  Google Scholar 

  • Cereghino JL, Cregg JM (2000) Heterologous protein expression in the methylotrophic yeast Pichia pastoris. FEMS Microbiol Rev 24:45–66

    Article  PubMed  CAS  Google Scholar 

  • Cesbron-Delauw MF (1994) Dense-granule organelles of Toxoplasma gondii: their role in the host–parasite relationship. Parasitol Today 10:293–296

    Article  PubMed  CAS  Google Scholar 

  • Cregg JM, Cereghino JL, Shi J, Higgins DR (2000) Recombinant protein expression in Pichia pastoris. Mol Biotechnol 16:23–52

    Article  PubMed  CAS  Google Scholar 

  • Darcy F, Maes P, Gras-Masse H (1992) Protection of mice and nude rats against toxoplasmosis by a multiple antigenic peptide construction derived from Toxoplasma gondii SAG1 antigen. J Immunol 149:3636–3641

    PubMed  CAS  Google Scholar 

  • Debard N, Buzoni-Gatel D, Bout D (1996) Intranasal immunization with SAG1 protein of Toxoplasma gondii in association with cholera toxin dramatically reduces development of cerebral cysts after oral infection. Infect Immun 64:2158–2166

    PubMed  CAS  Google Scholar 

  • Dubey JP, Thulliez P (1993) Persistence of tissue cysts in edible tissues of cattle fed T. gondii oocysts. Am J Vet Res 54:270–273

    PubMed  CAS  Google Scholar 

  • Frenkel JK (1988) Pathophysiology of toxoplasmosis. Parasitol Today 4:273–278

    Article  PubMed  CAS  Google Scholar 

  • Gazzinelli RT, Hakim FT, Hieny S, Shearer GM, Sher A (1991) Synergistic role of CD4+ and CD8+ T lymphocytes in IFN-gamma production and protective immunity induced by an attenuated Toxoplasma gondii vaccine. J Immunol 146:286–292

    PubMed  CAS  Google Scholar 

  • Godard I, Estaquier J, Zenner L, Bossus M, Auriault C, Darcy F, Gras-Masse H, Capron A (1994) Antigenicity and immunogenicity of P30-derived peptides in experimental models of toxoplasmosis. Mol Immunol 31:1353–1363

    Article  PubMed  CAS  Google Scholar 

  • Khan IA, Ely KH, Kasper LH (1991) A purified parasite antigen (p30) mediates CD8+ T cells immunity against fatal Toxoplasma gondii infection in mice. J Immunol 147:3501–3506

    PubMed  CAS  Google Scholar 

  • Kowalski JM, Parekh RN, Mao J, Wittrup KD (1998) Protein folding stability can determine the efficiency of escape from endoplasmic reticulum quality control. J Biol Chem 273:19453–19458

    Article  PubMed  CAS  Google Scholar 

  • Lunden A (1995) Immune response in sheep after immunization with Toxoplasma gondii antigens incorporated into iscoms. Vet Parasitol 56:23–35

    Article  PubMed  CAS  Google Scholar 

  • Macauley-Patrick S, Fazenda ML, McNeil B, Harvey LM (2005) Heterologous protein production using the Pichia pastoris expression system. Yeast 22:249–270

    Article  PubMed  CAS  Google Scholar 

  • Montoya JG, Liesenfeld O (2004) Toxoplasmosis. Lancet 363:1965–1976

    Article  PubMed  CAS  Google Scholar 

  • Murray A, Mercier C, Decoster A, Lecordier L, Capron A, Cesbron-Delauw MF (1993) Multiple B-cell epitopes in a recombinant GRA2 secreted antigen of Toxoplasma gondii. Appl Parasitol 34:235–244

    PubMed  CAS  Google Scholar 

  • Petersen E, Nielsen HV, Christiansen L, Spenter J (1998) Immunization with E. coli produced recombinant T. gondii SAG1 with alum as adjuvant protect mice against lethal infection with Toxoplasma gondii. Vaccine 16:1283–1289

    Article  PubMed  CAS  Google Scholar 

  • Pistoia V, Facchetti P, Ghiotto F, Cesbron-Delauw MF, Prigione I (1996) Characterization of human T cell clones specific for Toxoplasma gondii. Curr Top Microbiol Immunol 219:165–173

    PubMed  CAS  Google Scholar 

  • Prince JB, Araujo FG, Remington JS, Burg JL, Boothroyd JC, Sharma SD (1989) Cloning of cDNAs encoding a 28 kilodalton antigen of Toxoplasma gondii. Mol Biochem Parasitol 34:3–13

    Article  PubMed  CAS  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory, New York

    Google Scholar 

  • Sharma SD, Araujo FG, Remington JS (1984) Toxoplasma antigen isolated by affinity chromatography with monoclonal antibody protects mice against lethal infection with Toxoplasma gondii. J Immunol 133:2818–2820

    PubMed  CAS  Google Scholar 

  • Sreekrishna K, Brankamp RG, Kropp KE, Blankenship DT, Tsay JT, Smith PL, Wierschke JD, Subramaniam A, Birkenberger LA (1997) Strategies for optimal synthesis and secretion of heterologous proteins in the methylotrophic yeast Pichia pastoris. Gene 190:55–62

    Article  PubMed  CAS  Google Scholar 

  • Suzuki Y, Orellana MA, Schreiber RD, Remington JS (1988) Interferongamma: the major mediator of resistance against Toxoplasma gondii. Science 240:516–518

    Article  PubMed  CAS  Google Scholar 

  • Velge-Roussel F, Moretto M, Buzoni-Gatel D, Dimier-Poisson I, Ferrer M, Hoebeke J, Bout D (1997) Differences in immunological response to a T. gondii protein (SAG1) derived peptide between two strains of mice: effect on protection in T. gondii infection. Mol Immunol 34:1045–1053

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

This work was supported by the Natural Science Foundation of Shandong, China (No. Y2005C20) and by Fund Project of Medicine and Health Development Plan of Shandong, China (No. 2005HZ028). We declare that the experiments we performed comply with the current laws of China.

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  1. Department of Parasitology, School of Medicine, Shandong University, Jinan, 250012, People’s Republic of China

    Huaiyu Zhou, Qinmin Gu, Qunli Zhao, Jiaqin Zhang, Hua Cong, Ying Li & Shenyi He

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  1. Huaiyu Zhou
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  2. Qinmin Gu
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  3. Qunli Zhao
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  4. Jiaqin Zhang
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  5. Hua Cong
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  6. Ying Li
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  7. Shenyi He
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Correspondence to Qinmin Gu.

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Zhou, H., Gu, Q., Zhao, Q. et al. Toxoplasma gondii: expression and characterization of a recombinant protein containing SAG1 and GRA2 in Pichia pastoris . Parasitol Res 100, 829–835 (2007). https://doi.org/10.1007/s00436-006-0341-6

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  • DOI: https://doi.org/10.1007/s00436-006-0341-6

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