Interaction of Cholera Toxin with its Receptor the Monosialoganglioside GM1: A Fluorescence Study

  • Marc De Wolf
  • Géry Bastiaens
  • Albert Lagrou
  • Guido Van Dessel
  • Herwig Hilderson
  • Wilfried Dierick
Part of the NATO ASI Series book series (NSSA, volume 133)


Cholera toxin (CT) is an enterotoxin secreted by Vibrio cholerae producing its pathological effects by increasing the c-AMP level in intestinal epithelial cells1,2. It is an oligomeric protein (Mr ~ 84,000) composed of two structural and functional distinct subunits CT A and CT B (Mr ~ 29,000 and 55,000 respectively). CT B contains five identical polypeptide chains (Mr = 11,600), most likely arranged in a ring-like pentameric configuration and CT A consists of two non-identical polypeptide chains A or a-chain (Mr = 23,000) and A2 or γ-chain (Mr = 5,500) linked by a single disulfide bridge (for reviews see refs. 1, 3–5). CT A is synthesized as a single polypeptide chain which is “nicked” between the two cysteine residues by an extracellular bacterial protease. During this proteolysis two serine residues are removed at the COOH terminus of A1 6. The first event in the action of CT on cells is the rapid, irreversible binding to receptors on the cell surface. It is generally accepted that the receptor for the toxin is the mono-sialoganglioside GM1 (for reviews see refs. 3,4,7).


Lipid Bilayer Adenylate Cyclase Fluorescence Quenching Cholera Toxin Amino Acid Side Chain 
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.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    R.A. Finkelstein, Cholera, CRC Crit. Rev. Microbiol. 2:553–623 (1973).CrossRefGoogle Scholar
  2. 2.
    M. Field, Cholera toxin, adenylate cyclase, and the process of active secretion in the small intestine: The pathogenesis of diarrhea in cholera, in: “Physiology of Membrane Disorders”, T.E. Andreoli and J.F. Hoffman, eds., Plenum Publishing Corporation, New York, Chapter 41:877–898 (1978).CrossRefGoogle Scholar
  3. 3.
    J. Holmgren, Cholera toxin and the cell membrane, in: “Bacterial Toxins and Cell Membranes”, J. Jeljaszewick and T. Wadström, eds., Academic Press, New York, pp. 333–366 (1978).Google Scholar
  4. 4.
    V. Bennett, and P. Cuatrecasas, Cholera toxin: Membrane gangliosides and activation of adenylate cyclase, in: “The Specificity and Action of Animal, Bacterial and Plant Toxins”, P. Cuatrecasas and N.F. Greaves, eds., Chapman and Hall, London, pp. 3–66 (1977).Google Scholar
  5. 5.
    C.-Y. Lai, The chemistry and biology of cholera toxin, CRC Crit. Rev. Biochem. 9:171–206 (1980).PubMedCrossRefGoogle Scholar
  6. 6.
    L.K. Duffy, J.W. Peterson, and A. Kurosky, Isolation and characterization of a precursor form of the ‘A’ subunit of cholera toxin’ FEBS Letters 126:157–166 (1981).CrossRefGoogle Scholar
  7. 7.
    S. van Heijningen, Similarities in the action of different toxins, in: “Molecular Action of Toxins and Viruses”, C. Cohen and S. van Heijningen, eds., Elsevier Biochemical Press, Amsterdam, 6:169–190 (1982).Google Scholar
  8. 8.
    J. Moss, and M. Vaughan, Activation of adenylate cyclase by choleragen, Annu. Rev. Biochem. 48:581–600 (1979).PubMedCrossRefGoogle Scholar
  9. 9.
    G.L. Johnson, Cholera toxin action and the regulation of hormone-sensitive adenylate cyclase, in: “Molecular Actions of Toxins and Viruses”, P. Cohen and S. van Heijningen, eds., Elsevier, Amsterdam, pp. 33–49 (1982).Google Scholar
  10. 10.
    D. Cassel, and T. Pfeuffer, Mechanism of cholera toxin action: Covalent modification of the guanylnucleotide-binding protein of the adenylate cyclase system, Proc. Natl. Acad. Sci. USA 75:2669–2673 (1978).PubMedCrossRefGoogle Scholar
  11. 11.
    D. Cassel, and Z. Selinger, Mechanism of adenylate cyclase activation by cholera toxin: Inhibition of GTP hydrolysis at the regulatory site, Proc. Natl. Acad. Sci. USA 74:3307–3311 (1977).PubMedCrossRefGoogle Scholar
  12. 12.
    J.K. Northup, P.C. Sternweis, M.D. Smigel, L.S. Schleifer, E.M. Ross and A.G. Gilman, Purification of the regulatory component of adenylate cyclase, Proc. Natl. Acad. Sci. USA 77:6516–6520 (1980).PubMedCrossRefGoogle Scholar
  13. 13.
    T. Katada, and M. Ui, ADP ribosylation of the specific membrane protein of C6 cells by islet activaint protein associated with modification of adenylate cyclase activity, J. Biol. Chem. 257:7210–7216 (1982).PubMedGoogle Scholar
  14. 14.
    L. Olansky, G.A. Myers, S.L. Pohl, and E.L. Hewlett, Promotion of lipo-lysis in rat adipocytes by pertussis toxin: Reversal of endogenous inhibition, Proc. Natl. Acad. Sci. USA 80:6547–6551 (1983).PubMedCrossRefGoogle Scholar
  15. 15.
    A.G. Gilman, G Proteins and dual control of adenylate cyclase, Cell 36: 577–579 (1984).PubMedCrossRefGoogle Scholar
  16. 16.
    J.K. Northup, Overview of the guanine nucleotide regulatory protein systems, N and N., which regulate adenylate cyclase activity in plasma membranes, in: Molecular Mechanisms of Transmembrane Signalling, P. Cohen and M.D. Houslay, eds., Elsevier, Amsterdam, pp. 91–116 (1985).Google Scholar
  17. 17.
    R.A. Kahn, and A.G. Gilman, ADP-ribosylation of Gs promotes the dissociation of its a and β subunits, J. Biol. Chem. 259:6235–6240 (1984).PubMedGoogle Scholar
  18. 18.
    Z. Farfel, H.R. Kaslow, and H.R. Bourne, A regulatory component of adenylate cyclase is located on the inner surface of human erythrocyte membranes, Biochem. Biophys. Res. Commun. 90:1237–1241 (1979).PubMedCrossRefGoogle Scholar
  19. 19.
    H.R. Kaslow, G.L. Johnson, V.M. Brothers, and H.R. Bourne, A regulatory component of adenylate cyclase from human erythrocyte membranes, J. Biol. Chem. 255:3736–3741 (1980).PubMedGoogle Scholar
  20. 20.
    D.M. Gill, The arrangement of subunits in cholera toxin, Biochemistry 15:1242–1248 (1976).PubMedCrossRefGoogle Scholar
  21. 21.
    D.M. Gill, Seven toxic peptides that cross cell membranes, in: “Bacterial Toxins and Cell Membranes”, J. Jeljaswicz and T. Wadström, eds., Academic Press, London, pp. 291–332 (1978).Google Scholar
  22. 22.
    P.H. Fishman, Role of membrane gangliosides in the binding and action of bacterial toxins, J. Membrane Biol. 69:85–97 (1982).CrossRefGoogle Scholar
  23. 23.
    L.D. Kohn, E. Consiglio, M. De Wolf, E.F. Grollman, F.D. Ledley, G. Lee, and N.P. Morris, Thyrotropin receptors and gangliosides, Adv. Exp. Med. Biol. 125:487–503 (1980).PubMedGoogle Scholar
  24. 24.
    B.R. Mullin, S.M. Aloj, P.H. Fishman, G. Lee, L.D. Kohn, and R.O. Brady, Cholera toxin interactions with thyrotropin receptors on thyroid plasma membranes, Proc. Natl. Acad. Sci. USA 73:1679–1683 (1976).PubMedCrossRefGoogle Scholar
  25. 25.
    C.Y. Lai, Determination of the primary structure of cholera toxin B sub-unit, J. Biol. Chem. 252:7249–7256 (1977).PubMedGoogle Scholar
  26. 26.
    M.J.S. De Wolf, M. Fridkin, and L.D. Kohn, Tryptophan residues of cholera toxin and its A and B protomers: Intrinsic fluorescence and solute quenching upon interacting with the ganglioside GM1, oligo-GM1 or dan-sylated oligo-GM, J. Biol. Chem. 256:5489–5496 (1981a).PubMedGoogle Scholar
  27. 27.
    M.J.S. De Wolf, M. Fridkin, M. Epstein, and L.D. Kohn, Structure-function studies of cholera toxin and its A and B protomers. Modification of tryptophan residues, J. Biol. Chem. 256:5481–5488 (1981b).PubMedGoogle Scholar
  28. 28.
    M. De Wolf, G. Van Dessel, H.J. Hilderson, A. Lagrou, and W. Dierick, Acrylamide quenching of cholera toxin and its A and B protomers using steady state and dynamic fluorometry, Arch. internat. Physiol. Biochim. 91:B11-B13 (1983).Google Scholar
  29. 29.
    M. De Wolf, G. Van Dessel, A. Lagrou, H.J. Hilderson, and W. Dierick, Structural features of the binding site of cholera toxin inferred from fluorescence measurements, Biochim. Biophys. Acta, 832:165–174 (1985).PubMedCrossRefGoogle Scholar
  30. 30.
    M. Shinitzky, and R. Goldman, Fluorometric detection of histidine-tryptophan complexes in peptides and proteins, Eur. J. Biochem. 3:139–144 (1967).PubMedCrossRefGoogle Scholar
  31. 31.
    A. White, Effect of pH on fluorescence of tyrosine, tryptophan and related compounds, Biochem. J. 71:217–220 (1959).PubMedGoogle Scholar
  32. 32.
    I. Steinberg, Long-range nonradioactive transfer of electronic excitation energy in proteins and polypeptides, Annu. Rev. Biochem. 40: 83–114 (1971).PubMedCrossRefGoogle Scholar
  33. 33.
    J.J. Mekalanos, R.J. Collier, and W.R. Romig, Enzyme activity of cholera toxin. II. Relationships to proteolytic processing, disulfide bond reduction, and subunit composition, J. Biol. Chem. 254:5855–5861 (1979).PubMedGoogle Scholar
  34. 34.
    L.K. Duffy, and C.Y. Lai, Involvement of arginine residues in binding site of cholera toxin subunit B, Biochem. Biophys. Res. Commun. 91: 1005–1010 (1979).CrossRefGoogle Scholar
  35. 35.
    P.Y. Chou, and G.D. Fasman, Prediction of protein conformation, Biochemistry 13:222–244 (1974).PubMedCrossRefGoogle Scholar
  36. 36.
    J.D. Clements, and R.A. Finkelstein, Immunological cross-reactivity between a heat-labile enterotoxin(s) of Escherichia coli and subunits of Vibrio cholerae enterotoxin, Infect. Immun. 21:1036–1039 (1978).PubMedGoogle Scholar
  37. 37.
    S.L. Kunkel, and D.C. Robertson, Purification and chemical characterization of the heat-labile enterotoxin produced by enterotoxigenic Escherichia coli, Infect. Immun. 25:586–596 (1979).PubMedGoogle Scholar
  38. 38.
    W.S. Dallas, and S. Falkow, The molecular nature of heat-labile enterotoxin (LT) of Escherichia coli, Nature (London) 277:406–407 (1979).CrossRefGoogle Scholar
  39. 39.
    W.S. Dallas, and S. Falkow, Amino acid sequence homology between cholera toxin and Escherichia coli heat-labile toxin, Nature 288:499–501 (1980).PubMedCrossRefGoogle Scholar
  40. 40.
    J. Moss, J.C. Osborne, Jr., P.H. Fishman, S. Nakaya, and D.C. Robertson, Escherichia coli heat-labile enterotoxin ganglioside specificity and ADP-ribosyltransferase activity, J. Biol. Chem. 256:12861–12865 (1981).PubMedGoogle Scholar
  41. 41.
    F.D. Ledley, B.R. Mullin, G. Lee, S.M. Aloj, P.H. Fishman, L.T. Hunt, M.O. Dayhoff, and L.D. Kohn, Sequence similarity between cholera toxin and glycoprotein hormones: Implications for structure activity relationship and mechanism of action, Biochem. Biophys. Res. Commun. 69: 852–859 (1976).PubMedCrossRefGoogle Scholar
  42. 42.
    A. Kurosky, D.E. Markel, J.W. Peterson, and W.M. Fitch, Primary structure of cholera toxin ß-chain: A glycoprotein hormone analog ?, Science 195:299–301 (1977).PubMedCrossRefGoogle Scholar
  43. 43.
    M. De Wolf, G. Van Dessel, A. Lagrou, H.J. Hilderson, and W. Dierick, Hydrophobic fluorescence quenching of the subunits of cholera toxin: Possible relevance to lipid bilayer penetration, Arch, internat. Physiol. Biochim. 93:B136 (1985).Google Scholar
  44. 44.
    M. De Wolf, L.D. Kohn, H. Depauw, G. Van Dessel, H.J. Hilderson, A. Lagrou, and W. Dierick, Interaction of cholera toxin with thyroidal plasma membranes: Role of phospholipids, Arch, internat. Physiol. Biochim. 90:B110-B112 (1982).Google Scholar
  45. 45.
    W.H.J. Ward, P. Britton, and S. van Heijningen, The hydrophobicities of cholera toxin, tetanus toxin, and their components, Biochem. J. 199: 457–460 (1981).PubMedGoogle Scholar
  46. 46.
    M. Tomasi, and C. Montecucco, Lipid insertion of cholera toxin after binding to GM -containing liposomes, J. Biol. Chem. 256:11177–11181 (1981).PubMedGoogle Scholar
  47. 47.
    B.J. Wisnieski, and J.S. Bramhall, Photolabelling of cholera toxin sub-units during membrane penetration, Nature 289:319–321 (1981).PubMedCrossRefGoogle Scholar
  48. 48.
    K.C. Joseph, S.U. Kim, A. Stieber, and N.K. Gonatas, Endocytosis of cholera toxin into neuronal GERL, Proc. Natl. Acad. Sci. USA 75:2815–2819 (1978).PubMedCrossRefGoogle Scholar
  49. 49.
    P.P. Chang, P.H. Fishman, N. Ohtomo, and J. Moss, Degradation of choleragen bound to cultured human fibroblasts and mouse neuroblastoma cells, J. Biol. Chem. 258:426–430 (1983).PubMedGoogle Scholar
  50. 50.
    M.D. Houslay, and K.R.F. Elliott, Is the receptor-mediated endocytosis of cholera toxin a pre-requisite for its activation of adenylate cyclase in intact rat hepatocytes, FEBS Letters 128:289–292 (1981).PubMedCrossRefGoogle Scholar
  51. 51.
    J. Hagman, and P.H. Fishman, Inhibitors of protein synthesis block action of cholera toxin, Biochem. Biophys. Res. Commun. 98:677–684 (1981).CrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1987

Authors and Affiliations

  • Marc De Wolf
    • 1
  • Géry Bastiaens
    • 1
  • Albert Lagrou
    • 1
  • Guido Van Dessel
    • 1
  • Herwig Hilderson
    • 1
  • Wilfried Dierick
    • 1
  1. 1.RUCA-Laboratory for Human Biochemistry and UIA-Laboratory for Pathological BiochemistryUniversity of AntwerpAntwerpBelgium

Personalised recommendations