Techniques for Conjugation of Synthetic Peptides to Carrier Molecules

  • J. Mark Carter
Part of the Methods in Molecular Biology book series (MIMB, volume 36)


There are two common purposes for conjugation of peptides. The most common is induction of humoral immunity. This is the production of antibodies capable of binding to the peptide immunogen. The antibodies are elaborated by plasma cells, which are terminally differentiated B-lymphocytes. However, in order for immunity to be successfully induced in a secondary anamnestic response, the immunogen must also react with T-lymphocytes. Many peptides contain B-cell epitopes, but not T-cell epitopes. In immunological terms, these peptides and other such molecules are called haptens. Coupling these molecules to a large carrier protein containing T-cell epitopes allows the induction of a B-cell response to the entire immunogen, including the peptide (1). New synthetic peptides offer promise as vaccines.


Disulfide Bond Carrier Protein Keyhole Limpet Hemocyanin Free Thiol Conjugation Reaction 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


  1. 1.
    Tanaka, T., Slamon, D. J., and Line, M J. (1985) Efficient generation of antibodies to oncogene proteins by using synthetic peptide antigens. Proc. Natl. Acad. Sci. USA 82, 3400–3404.PubMedCrossRefGoogle Scholar
  2. 2.
    Tam, J. P. and Zavala, F. (1989) Multiple antigen peptides. J. Immunol. Meth. 124, 53–61.CrossRefGoogle Scholar
  3. 3.
    VanRegenmortel, M. H. V., Briand, J. P., Muller, S., and Plaue, S. Luboratory Techniques in Biochemistry ana Molecular Biology, vol 19 (Burdon, R. H. and Van Knippenberg, P. H., eds.), Elsevier, Amsterdam.Google Scholar
  4. 4.
    Dryberg, T. and Oldstone, M. B. A. (1986) Peptides as antigens. J. Exp. Med. 164, 1344–1349.CrossRefGoogle Scholar
  5. 5.
    Ponsati, B., Giraldt, E., and Andreu, D. (1989) A synthetic strategy for simultaneous purification-conjugation of antigenic peptides. Analytical Biochem. 181, 389–395.CrossRefGoogle Scholar
  6. 6.
    Satterthwait, A. C., Arrhenius, T., Hagopian, R. A., Zavala, F., Nussenzweig, V., and Lerner, R. A. (1988) Conformational restriction of peptidyl immunogens with covalent replacements for the hydrogen bond. Vaccine 6, 99–103.PubMedCrossRefGoogle Scholar
  7. 7.
    Peeters, J. M., Hazendonk, T. G., Beuvery, E. C., and Tesser, G. I. (1989) Comparison of four bifunctional reagents for coupling peptides to proteins and the effect of the three moieties on the immunogenicity of the conjugates. J. Immunol. Methods, 120, 133–143.PubMedCrossRefGoogle Scholar
  8. 8.
    Ruegg, U. T. and Rudinger, J. (1977) Reductive cleavage of cystine disulfides with tributylphosphine. Meth. Enzymol, 47, 11l–l16.Google Scholar
  9. 9.
    Atassi, M. Z. and Habeeb, A. F. S. A. (1972) Reactions of proteins with citraconic anhydride. Meth. Enzymol. 25, 546.CrossRefGoogle Scholar
  10. 10.
    Schaaper, W. M. M., Lankohof, H., Pujik, W. C., and Meleon, R. H. (1989) Manipulation of antipeptide immune response by varying the coupling of the peptide with the carrier protein. Mol. Immunol. 26, 81–86.PubMedCrossRefGoogle Scholar
  11. 11.
    Reichlin, M. (1980) Use of glutaraldehyde as a coupling agent for proteins and peptides. Meth. Enzymol. 70, 159–165.PubMedCrossRefGoogle Scholar
  12. 12.
    Kirkeby, S., Jakobsen, P., and Moe, D. (1987) Glutaraldehyde—pure and impure. A spectroscopic investigation of two commercial glutaraldehyde solutions and their reaction products with amino acids. Analyt. Lett. 20(2), 303–315.Google Scholar
  13. 13.
    Baron, M. H. and Baltimore, D. (1982) Antibodres against the chemically synthesized genome-linked protein of polio virus react with native virus-specific proteins. Cell 28, 395–404.PubMedCrossRefGoogle Scholar
  14. 14.
    Bauminger, S. and Wilchek, M(1980) The use of carbodiimides in the preparation of immunizing comugates. Meth. Enzymol. 70, 151–159.PubMedCrossRefGoogle Scholar
  15. 15.
    Liu, F.-T, Zinnecker, M., Hamaoka, T., and Katz, D H. (1979) New procedures for preparation and isolation of conjugates of proteins and a synthetic copolymer of D-amino acids and immunochemical characterization of such conmgates. Biochem 18(4), 690–697.CrossRefGoogle Scholar
  16. 16.
    Lerner, R. A., Green, N, Alexander, H., Liu, F-T., Sutcliffe, J. G, and Shinnick, T M. (1981) Chemically synthesized peptides predicted from the nucleotide sequence of the Hepatitis B virus genome elicit antibodies reactive with the native envelope protein of Dane particles. Proc Nat1 Acad. Sci. USA 78, 3403–3407.CrossRefGoogle Scholar
  17. 17.
    Kolodny, N and Robey, F. A. (1990) Conjugation of synthetic peptides to proteins: quantitation from S-carboxymethylcysteine released upon acid hydrolysis. Anal. Biochem. 187, 136–140PubMedCrossRefGoogle Scholar
  18. 18.
    Lmdler, W. and Robey, F. A (1987) Automated synthesis and use of N-chloro-acetyl-modified peptides for the preparation of synthetic pepttde polymers and peptide-protein tmmunogens. Int. J. Peptide Protem Res. 30, 794–800Google Scholar
  19. 19.
    Inman, J. K., Highet, P. F., Kolodny, N., and Robey, F A. (1991) Synthesis of N-alpha-(tert-butoxycarbonyl)-N-epsilon-[(N-bromoacetyl)-beta-alanyl]-L-lysine tts use in peptide synthesis for placing a bromoacetyl cross-linking function at any desired sequence position. Bioconjug. Chem. 2, 458–463.PubMedCrossRefGoogle Scholar
  20. 20.
    Weir, D. M. (ed.) (1986) Handbook Of Experimental Immunology, vol. 1, Blackwell Scientific, Oxford, p 20.14.Google Scholar
  21. 21.
    Gordon, R D, Fteles, W. E., Schotland, D. L., Hogue-Angelettt, R, and Barchi, R. L. (1987) Topographical localization of the C-terminal region of the voltage-dependent sodium channel from Electrophorus Electricus using antibodies raised against a synthetic peptide. Proc. Natl. Acad. Sci. USA 84, 308–312.PubMedCrossRefGoogle Scholar
  22. 22.
    Carlsson, J., Drevin, H., and Axen, R. (1978) Protein thiolation and reversible protem-protein conjugation. Biochem. J 173, 723–737.PubMedGoogle Scholar
  23. 23.
    Tam, J. P. (1988) Synthetic peptide vaccine design: synthesis and properties of a high-density multiple anttgemc pepttde system. Proc. Natl. Acad. Sci. USA 85, 5409–5413.PubMedCrossRefGoogle Scholar
  24. 24.
    Tam, J. P. and Lu, Y. A. (1989) Vaccine engineering: enhancement of immu-nogemcity of synthetic peptide vaccmes for Hepatitis in chemically defined models consisting of T-and B-cell epitopes. Proc. Natl. Acad Sci. USA 86, 9084–9088.PubMedCrossRefGoogle Scholar
  25. 25.
    Borras-Cuesta, F., Petit-Camurdan, A., and Fedon, Y. (1987) Engineermg of immunogenic peptides by co-linear synthesis of determinants recognized by B and T cells. Eur. J. Immunol. 17, 1213–1215, also Borras-Cuesta, F., Fedon, Y., and Petit-Camurdan, A. (1988) Enhancement of peptide immunogenicity by lmear polymerization. Eur. J Zmmunof. l8, 199–202.PubMedCrossRefGoogle Scholar
  26. 26.
    Good, M. F., Maloy, W. L., Lunde, M. N., Margalit, H., Cornette, J. L., Smith, G. L., Moss, B., Miller, L. H., and Berzofsky, J. A. (1987) Construction of a synthetic immunogen: use of a new T-helper epitope on malaria circumsporozoite protein. Science 235, 1059–1062.PubMedCrossRefGoogle Scholar
  27. 27.
    LeClerc, C., Przewlocki, G., Schutze, M. P., and Chedid, L. (1987) A synthetic vaccine constructed by copolymerization of B and T cell determinants. Eur. J. Immunol. 17, 269–273PubMedCrossRefGoogle Scholar
  28. 28.
    Patarroyo, M. E., Amador, R., Clavijo, P., Moreno, A., Guzman, F., Romero, P., Tascon, R., France, A., Murillo, L. A., Ponton, G., and Trujillo, G. (1988) A synthetic vaccine protects humans against challenge with asexual blood stages of Plus-modiumfalciparum malaria. Nature 332, 158–161.PubMedCrossRefGoogle Scholar
  29. 29.
    Alving, R., Richards, R. L., Moss, J., Alving, L. I., Clements, J. D., Shiba, T., Kotani, S., Wirtz, R. A., and Hockmeyer, W. T. (1986) Effectiveness of liposomes as potential carriers of vaccines applications to cholera toxin and human malaria sporozoite antigen Vaccine 4, 166–172.PubMedCrossRefGoogle Scholar
  30. 30.
    Hui, G. S. N., Chang, S. P., Gibson, H., Hashimoto, A., Hashiro, C, Barr, P J., and Kotani, S. (1991) Influence of adJuvants on the antibody specificity to the Plasmodiumfalciparum major merozoite surface protein, gp195. J. lmmunol. 147, 3935–3941Google Scholar
  31. 31.
    Deres, K, Schild, H., Weissmuller, K. H., and Jung, G. (1989) In viva priming of virus-specific cytotoxic T lymphocytes with synthetic lipopeptide vaccine. Nature 342, 561–564.PubMedCrossRefGoogle Scholar
  32. 32.
    Weissmuller, K. G., Jung, G., and Hess, G. (1989) Novel low-molecular-weight synthetic vaccine against foot-and-mouth disease containing a potent B-Cell and macrophage activator. Vaccine 7, 29–33.CrossRefGoogle Scholar
  33. 33.
    Hopp, T. P. (1984) Immunogenicity of a synthetic HBsAg peptide enhancement by conjugation to a fatty acid carrier, Molecular Immunol. 21, 13–16.CrossRefGoogle Scholar
  34. 34.
    Lyon, J. A., Geller, R. H., Haynes, J. D, Chulay, J. D., and Weber, J L (1986) Epitope map and processing scheme for the 195,000 dalton surface glycoprotein of Plasmodiumfulciparum merozoite deduced from cloned overlapping segments of the gene. Proc. Natl. Acad. SCL USA 83, 2989–2993.CrossRefGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 1994

Authors and Affiliations

  • J. Mark Carter
    • 1
  1. 1.CytogenPrinceton

Personalised recommendations