Chimeric Proteins

  • Riny Janssen
  • Jan Tommassen
Part of the Methods in Molecular Medicine™ book series (MIMM, volume 4)

Abstract

Most conventional vaccines consist of killed organisms or purified antigemc components of pathogens. However, this approach of vaccine development has several limitations. Large-scale growth of pathogens may be difficult to achieve and is not completely free of risk. Furthermore, purification of relevant antigenic components from such pathogens may be expensive and time-consuming. Another approach is the use of synthetic peptides corresponding to immunogenic epitopes of pathogens. However, peptides are not immunogenic by themselves and need to be coupled to a carrier (see  Chapter 6). Recent developments in molecular biology have opened up new possibilities for the production of vaccines and serum diagnostics. Protein antigens derived from pathogens can be expressed in attenuated bacteria, such as Salmonella aroA mutants (1 and see  Chapter 4), which can be used as live oral vaccines. Such Salmonella strains have been shown to induce humoral, cellular, and mucosal immune responses in animals and humans (2, 3, 4). High-level expression of foreign antigens can be achieved in Salmonella using strong promoters (5). However, high-level expression of heterologous proteins can be deleterious or even lethal to bacteria. On the other hand, the foreign proteins may be rapidly degraded in the heterologous host. It might be more convenient, therefore, to create chimeric proteins by inserting well-defined epitopes of antigens derived from pathogens into bacterial carrier proteins, which can be expressed at high levels.

Keywords

Phenol Codon EDTA Agarose Chloroform 

References

  1. 1.
    Hoiseth, S.K. and Stocker, B. A. D. (1981) Aromatic-dependent SaZmonelZa typhzmurium are non-virulent and effective as live vaccines. Nature 291, 238,239.CrossRefGoogle Scholar
  2. 2.
    Dougan, G., Sellwood, R., Maskell, D., Sweeney, K., Liew, F. Y., Beesely, J., and Hormaeche, C. (1986) In viva properties of a cloned K88 adherence antigen determinant. Infect. Immun. 52, 344–347.PubMedGoogle Scholar
  3. 3.
    Brown, A., Hormaeche, C. E., Dennarco de Hormaeche, R., Winther, M., Dougan, G., Maskell, D. J., and Stocker, B. A. D. (1987) An attenuated aroA SuZmoneZZu typhimurium vaccine elicits humoral and cellular immunity to cloned β-galactosi-dase in mice. J. Infect. Du. 155, 86–92.Google Scholar
  4. 4.
    Cebra, J. J., Gearhart, P. J., Kamat, R., Robertson, S.M., and Tseng, J. (1976) Origin and differentiation of lymphocytes involved in secretory IgA response. Cold Spring Harbor Symp. Quant. Biol. 41, 201–215.Google Scholar
  5. 5.
    Chatfield, S. N., Charles, I. G., Makoff, A. J., Oxer, M. D., Dougan, G, Pickard, D., Slater, D., and Fanweather, N. F. (1992) Use of the nirB promoter to direct the stable expression of heterologous antigens in Salmonella oral vaccine strains: development of a single-dose oral tetanus vaccine. Biotechnology 10, 888–892.PubMedCrossRefGoogle Scholar
  6. 6.
    Janssen, R. and Tommassen, J. (1994) PhoE protein as a carrier for foreign epitopes. Int. Rev. Immunol. 11, 113–121.PubMedCrossRefGoogle Scholar
  7. 7.
    Hofnung, M. and Charbit, A. (1992) Expression antigens as recombinant proteins, in Structure of dntzgens, vol 2. (Van Regenmortel, M. H. V., ed.), Telford, NJGoogle Scholar
  8. 8.
    Stanley, K.K. and Luzio, J P. (1984) Construction of a new family of high efficiency bacterial expression vectors: identification of cDNA clones coding for human liver proteins. EMBO J 3, 1429–1434PubMedGoogle Scholar
  9. 9.
    Newton, S. M. C., Jacob, C.O. and Stocker, B. A. D (1989) Immune response to cholera toxin epitope inserted in Salmonella flagellin. Science 244, 70–72.PubMedCrossRefGoogle Scholar
  10. 10.
    Van Die, I., Wauben, M., van Megen, I., Bergmans, H., Rtegman, N., Hoekstra, W., Pouwels, P., and Enger-Valk, E. L. (1988) Genetic mampulation of major β-fimbrial subunits and consequences for formation of fimbriae. J. Bacterial. 170, 5870–5876.Google Scholar
  11. 11.
    Agterberg, M., Adriaanse, H., Balteling, S., van Maanen, K., and Tommassen, J (1990) Protection of guinea-pigs against foot-and-mouth disease virus by immunization with a PhoE-FMDV hybrid protein Vaccine 8, 438–440.PubMedCrossRefGoogle Scholar
  12. 12.
    Charbit, A., Ronco, J., Michel, V., Werts, C., and Hofnung, M. (1991) Permissive sates and topology of an outer membrane protein with a reporter eprtope. J. Bactenol. 173, 262–275Google Scholar
  13. 13.
    Janssen, R., Wauben, M., van der Zee, R., de Gast, M., and Tommassen, J. (1994) Influence of ammo acids of a carrier protein flanking an inserted T cell determinant on T cell stimulatron. Int. Immunol. 6, 1187–1193.PubMedCrossRefGoogle Scholar
  14. 14.
    Agterberg, M., Benz, R., and Tommassen, J. (1987) Insertion mutagenesis on a cell-surface-exposed region of outer membrane protein PhoE of Escherichia cob K-12. Eur. J. Brochem. 169, 65–71.CrossRefGoogle Scholar
  15. 15.
    Charbit, A., Boulain, J. C., Ryter, A., and Hofnung, M. (1986) Probing the topology of a bacterial membrane protein by genetic insertion of a foreign epitope, expression at the cell surface. EMBOJ. 5, 3029–3037.Google Scholar
  16. 16.
    Duplay, P, Szmelcman, S., Bedouelle, H., and Hofnung, M (1987) Silent and functional changes in the periplasmic maltose-bmding protein of Escherichla coh K12.I. Transport of maltose. J. Mol. Biol. 194, 663–673.PubMedCrossRefGoogle Scholar
  17. 17.
    Ikemura, T (1981) Correlation between the abundance of Escherichla coli transfer RNAs and the occurrence of the respective codons in Its protein genes: a proposal for a synonymous codon choice that is optnnal for the E. colz translatronal system. J. Mol. Biol. 151, 389–409.PubMedCrossRefGoogle Scholar
  18. 18.
    Tommassen, J., van Tol, H, and Lugtenberg, B. (1983) The ultimate localization of an outer membrane protein of Escherzchia cob K-12 is not determined by the signal sequence. EMBOJ. 2, 1275–1279Google Scholar
  19. 19.
    Korteland, J., Overbeeke, N., de Graaff, P., Overduin, P, and Lugtenberg, B (1985) Role of the Arg158 residue of the outer membrane PhoE pore protein of Escherrchia coli K-12 in bacteriophage TC45 recognitron and in channel charactenstrcs. Eur. J Blochem 152, 691–697CrossRefGoogle Scholar
  20. 20.
    Boulam, J. C., Charbrt, A., and Hofnung, M. (1986) Mutagenesis by random linker insertion into the ZamB gene of Escherichza colz K12. Mol. Gen. Genet. 205, 339–348.CrossRefGoogle Scholar

Copyright information

© Humana Press Inc, Totowa, NJ 1996

Authors and Affiliations

  • Riny Janssen
    • 1
  • Jan Tommassen
    • 1
  1. 1.Department of Molecular Cell BiologyUtrecht UniversityUtrechtThe Netherlands

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