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. 1980 Apr;77(4):1986–1990. doi: 10.1073/pnas.77.4.1986

Binding of diphtheria toxin to phospholipids in liposomes

Carl R Alving *, Barbara H Iglewski †, Katharine A Urban *, Joel Moss ‡, Roberta L Richards *, Jerald C Sadoff §
PMCID: PMC348635  PMID: 6929533

Abstract

Diphtheria toxin bound to the phosphate portion of some, but not all, phospholipids in liposomes. Liposomes consisting of dimyristoyl phosphatidylcholine and cholesterol did not bind toxin. Addition of 20 mol% (compared to dimyristoyl phosphatidylcholine) of dipalmitoyl phosphatidic acid, dicetyl phosphate, phosphatidylinositol phosphate, cardiolipin, or phosphatidylserine in the liposomes resulted in substantial binding of toxin. Inclusion of phosphatidylinositol in dimyristol phosphatidylcholine/cholesterol liposomes did not result in toxin binding. The calcium salt of dipalmitoyl phosphatidic acid was more effective than the sodium salt, and the highest level of binding occurred with liposomes consisting only of dipalmitoyl phosphatidic acid (calcium salt) and cholesterol. Binding of toxin to liposomes was dependent on pH, and the pattern of pH dependence varied with liposomes having different compositions. Incubation of diphtheria toxin with liposomes containing dicetyl phosphate resulted in maximal binding at pH 3.6, whereas binding to liposomes containing phosphatidylinositol phosphate was maximal above pH 7. Toxin did not bind to liposomes containing 20 mol% of a free fatty acid (palmitic acid) or a sulfated lipid (3-sulfogalactosylceramide). Toxin binding to dicetyl phosphate or phosphatidylinositol phosphate was inhibited by UTP, ATP, phosphocholine, or p-nitrophenyl phosphate, but not by uracil. We conclude that (a) diphtheria toxin binds specifically to the phosphate portion of certain phospholipids, (b) binding to phospholipids in liposomes is dependent on pH, but is not due only to electrostatic interaction, and (c) binding may be strongly influenced by the composition of adjacent phospholipids that do not bind toxin. We propose that a minor membrane phospholipid (such as phosphatidylinositol phosphate or phosphatidic acid), or that some other phosphorylated membrane molecule (such as a phosphoprotein) may be important in the initial binding of diphtheria toxin to cells.

Keywords: membranes, receptors

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1986

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Alving C. R., Conrad D. H., Gockerman J. P., Gibbs M. B., Wirtz G. H. Vitamin A in liposomes. Inhibition of complement binding and alteration of membrane structure. Biochim Biophys Acta. 1975 Jun 25;394(2):157–165. doi: 10.1016/0005-2736(75)90253-9. [DOI] [PubMed] [Google Scholar]
  2. Alving C. R., Fowble J. W., Joseph K. C. Comparative properties of four galactosyl lipids as antigens in liposomes. Immunochemistry. 1974 Aug;11(8):475–481. doi: 10.1016/0019-2791(74)90118-9. [DOI] [PubMed] [Google Scholar]
  3. Alving C. R., Habig W. H., Urban K. A., Hardegree M. C. Cholesterol-dependent tetanolysin damage to liposomes. Biochim Biophys Acta. 1979 Feb 20;551(1):224–228. doi: 10.1016/0005-2736(79)90368-7. [DOI] [PubMed] [Google Scholar]
  4. Alving C. R., Kinsky S. C. The preparation and properties of liposomes in the LA and LAC states. Immunochemistry. 1971 Apr;8(4):325–343. doi: 10.1016/0019-2791(71)90155-8. [DOI] [PubMed] [Google Scholar]
  5. Alving C. R., Richards R. L. Immune reactivities of antibodies against glycolipids--I. Properties of anti-galactocerebroside antibodies purified by a novel technique of affinity binding to liposomes. Immunochemistry. 1977 May;14(5):373–381. [PubMed] [Google Scholar]
  6. Bernheimer A. W., Avigad L. S. Properties of a toxin from the sea anemone Stoichacis helianthus, including specific binding to sphingomyelin. Proc Natl Acad Sci U S A. 1976 Feb;73(2):467–471. doi: 10.1073/pnas.73.2.467. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bernheimer A. W. Cytolytic toxins of bacterial origin. The nature and properties of cytolytic proteins are discussed with emphasis on staphylococcal alpha-toxin. Science. 1968 Feb 23;159(3817):847–851. doi: 10.1126/science.159.3817.847. [DOI] [PubMed] [Google Scholar]
  8. Buckley J. T. Coisolation of glycophorin A and polyphosphoinositides from human erythrocyte membranes. Can J Biochem. 1978 May;56(5):349–351. doi: 10.1139/o78-055. [DOI] [PubMed] [Google Scholar]
  9. Chang T., Neville D. M., Jr Demonstration of diphtheria toxin receptors on surface membranes from both toxin-sensitive and toxin-resistant species. J Biol Chem. 1978 Oct 10;253(19):6866–6871. [PubMed] [Google Scholar]
  10. Clowes A. W., Cherry R. J., Chapman D. Physical effects of tetanus toxin on model membranes containing ganglioside. J Mol Biol. 1972 Jun 14;67(1):49–57. doi: 10.1016/0022-2836(72)90385-3. [DOI] [PubMed] [Google Scholar]
  11. Cuatrecasas P. Interaction of Vibrio cholerae enterotoxin with cell membranes. Biochemistry. 1973 Aug 28;12(18):3547–3558. doi: 10.1021/bi00742a031. [DOI] [PubMed] [Google Scholar]
  12. Cukor G., Solotorovsky M., Kuchler R. J. Biological activity of heated diphtheria toxin. J Bacteriol. 1973 Jul;115(1):277–283. doi: 10.1128/jb.115.1.277-283.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dimpfel W., Huang R. T., Habermann E. Gangliosides in nervous tissue cultures and binding of 125I-labelled tetanus toxin, a neuronal marker. J Neurochem. 1977 Aug;29(2):329–334. doi: 10.1111/j.1471-4159.1977.tb09626.x. [DOI] [PubMed] [Google Scholar]
  14. Donta S. T., Viner J. P. Inhibition of the steroidogenic effects of cholera and heat-labile Escherichia coli enterotoxins by GM1 ganglioside: evidence for a similar receptor site for the two toxins. Infect Immun. 1975 May;11(5):982–985. doi: 10.1128/iai.11.5.982-985.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Draper R. K., Chin D., Simon M. I. Diphtheria toxin has the properties of a lectin. Proc Natl Acad Sci U S A. 1978 Jan;75(1):261–265. doi: 10.1073/pnas.75.1.261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Duncan J. L., Groman N. B. Activity of diphtheria toxin. II. Early events in the intoxication of HeLa cells. J Bacteriol. 1969 Jun;98(3):963–969. doi: 10.1128/jb.98.3.963-969.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Holmgren J., Lönnroth I., Svennerholm L. Tissue receptor for cholera exotoxin: postulated structure from studies with GM1 ganglioside and related glycolipids. Infect Immun. 1973 Aug;8(2):208–214. doi: 10.1128/iai.8.2.208-214.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Imai M., Inoue K. The mechanism on the action of prymnesium toxin on membranes. Biochim Biophys Acta. 1974 Jun 29;352(3):344–348. doi: 10.1016/0005-2736(74)90226-0. [DOI] [PubMed] [Google Scholar]
  19. Kinsky S. C. Preparation of liposomes and a spectrophotometric assay for release of trapped glucose marker. Methods Enzymol. 1974;32:501–513. doi: 10.1016/0076-6879(74)32050-2. [DOI] [PubMed] [Google Scholar]
  20. Middlebrook J. L., Dorland R. B., Leppla S. H. Association of diphtheria toxin with Vero cells. Demonstration of a receptor. J Biol Chem. 1978 Oct 25;253(20):7325–7330. [PubMed] [Google Scholar]
  21. Middlebrook J. L., Dorland R. B., Leppla S. H. Effects of lectins on the interaction of diphtheria toxin with mammalian cells. Exp Cell Res. 1979 Jun;121(1):95–101. doi: 10.1016/0014-4827(79)90448-8. [DOI] [PubMed] [Google Scholar]
  22. Middlebrook J. L., Dorland R. B. Protection of mammalian cells from diphtheria toxin by exogenous nucleotides. Can J Microbiol. 1979 Mar;25(3):285–290. doi: 10.1139/m79-046. [DOI] [PubMed] [Google Scholar]
  23. Moehring T. J., Crispell J. P. Enzyme treatment of KB cells: the altered effect of diphtheria toxin. Biochem Biophys Res Commun. 1974 Oct 23;60(4):1446–1452. doi: 10.1016/0006-291x(74)90360-x. [DOI] [PubMed] [Google Scholar]
  24. Mooney J. J., Dalrymple J. M., Alving C. R., Russell P. K. Interaction of Sindbis virus with liposomal model membranes. J Virol. 1975 Feb;15(2):225–231. doi: 10.1128/jvi.15.2.225-231.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Moss J., Fishman P. H., Richards R. L., Alving C. R., Vaughan M., Brady R. O. Choleragen-mediated release of trapped glucose from liposomes containing ganglioside GM1. Proc Natl Acad Sci U S A. 1976 Oct;73(10):3480–3483. doi: 10.1073/pnas.73.10.3480. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Pappenheimer A. M., Jr Diphtheria toxin. Annu Rev Biochem. 1977;46:69–94. doi: 10.1146/annurev.bi.46.070177.000441. [DOI] [PubMed] [Google Scholar]
  27. Proia R. L., Hart D. A., Eidels L. Interaction of diphtheria toxin with phosphorylated molecules. Infect Immun. 1979 Dec;26(3):942–948. doi: 10.1128/iai.26.3.942-948.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Proia R. L., Hart D. A., Holmes R. K., Holmes K. V., Eidels L. Immunoprecipitation and partial characterization of diphtheria toxin-binding glycoproteins from surface of guinea pig cells. Proc Natl Acad Sci U S A. 1979 Feb;76(2):685–689. doi: 10.1073/pnas.76.2.685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Rothman J. E., Lenard J. Membrane asymmetry. Science. 1977 Feb 25;195(4280):743–753. doi: 10.1126/science.402030. [DOI] [PubMed] [Google Scholar]
  30. Simpson L. L., Rapport M. M. The binding of botulinum toxin to membrane lipids: sphingolipids, steroids and fatty acids. J Neurochem. 1971 Sep;18(9):1751–1759. doi: 10.1111/j.1471-4159.1971.tb03750.x. [DOI] [PubMed] [Google Scholar]
  31. Van Zoelen E. J., Zwaal R. F., Reuvers R. A., Demel R. A., Van Deenen L. L. Evidence for the preferential interaction of glycophorin with negatively charged phospholipids. Biochim Biophys Acta. 1977 Feb 4;464(3):482–492. doi: 10.1016/0005-2736(77)90024-4. [DOI] [PubMed] [Google Scholar]

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