1887

Abstract

The 16S toxin and one subcomponent of haemagglutinin (HA), designated HA1, were purified from a type D culture of by a newly established procedure, and their HA activities as well as that of purified type C 16S toxin were characterized. SDS-PAGE analysis indicated that the free HA1 forms a polymer with a molecular mass of approximately 200 kDa. Type C and D 16S toxins agglutinated human erythrocytes in the same manner. Their HA titres were dramatically reduced by employing erythrocytes that had been previously treated with neuraminidase, papain or proteinase K, and were inhibited by the addition of -acetylneuraminic acid to the reaction mixtures. In a direct-binding test to glycolipids such as SPG (NeuAcα2-3Galβ1-4GlcNAcβ1-3Galβ1-4Glcβ1-Cer) and GM3 (NeuAcα2-3Galβ1-4Glcβ1-Cer), and glycoproteins such as glycophorin A and/or B prepared from the erythrocytes, both toxins bound to sialylglycolipids and sialoglycoproteins, but bound to neither neutral glycolipids nor asialoglycoproteins. On the basis of these results, it was concluded that type C and D 16S toxins bind to erythrocytes through -acetylneuraminic acid. HA1 showed no haemagglutination activity, although it did bind to sialylglycolipids. We therefore speculate that binding to glycoproteins rather than to glycolipids may be important in causing haemagglutination by type C and D 16S toxins.

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1999-09-01
2024-03-28
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References

  1. Alberts B., Bray D., Lewis J., Raff M., Roberts K., Watson J. D. 1995 The Molecular Biology of the Cell, 3rd edn. New York: Garland Publishing;
    [Google Scholar]
  2. Balding P., Gold E. R., Boroff D. A., Roberts T. A. 1973; Observations on receptor specific proteins. II. Hemagglutination and hemagglutination-inhibition reactions of Clostridium botulinum types A, C, D, and E hemagglutinins. Immunology 25:773–782
    [Google Scholar]
  3. Fu F.-N., Sharma S. K., Singh B. R. 1998; A protease-resistant novel hemagglutinin purified from type A Clostridium botulinum.. J Protein Chem 17:53–60 [CrossRef]
    [Google Scholar]
  4. Fujinaga Y., Inoue K., Shimazaki S.8 other authors 1994; Molecular construction of Clostridium botulinum type C progenitor toxin and its gene organization. Biochem Biophys Res Commun 205:1291–1298 [CrossRef]
    [Google Scholar]
  5. Fujinaga Y., Inoue K., Watanabe S., Yokota K., Hirai Y., Nagamachi E., Oguma K. 1997; The haemagglutinin of Clostridium botulinum type C progenitor toxin plays an essential role in binding of toxin to the epitherial cells of guinea pig small intestine, leading to the efficient absorption of the toxin. Microbiology 143:3841–3847 [CrossRef]
    [Google Scholar]
  6. Gasa S., Makita A., Kinoshita Y. 1983; Further study of the chemical structure of the equine erythrocyte hematoside containing O-acetyl ester. J Biol Chem 258:876–881
    [Google Scholar]
  7. Hauser D., Eklund M. W., Boquet P., Popoff M. R. 1994; Organization of the botulinum neurotoxin C1 gene and its associated non-toxic protein genes in Clostridium botulinum C 468. Mol Gen Genet 243:631–640
    [Google Scholar]
  8. Hirano H., Watanabe T. 1990; Microsequencing of proteins electrotransferred onto immobilizing matrices from polyacrylamide gel electrophoresis: application to an insoluble protein. Electrophoresis 11:573–580 [CrossRef]
    [Google Scholar]
  9. Inoue K., Fujinaga Y., Watanabe T., Ohyama T., Takeshi K., Moriishi K., Nakajima H., Inoue K., Oguma K. 1996; Molecular composition of Clostridium botulinum type A progenitor toxins. Infect Immun 64:1589–1594
    [Google Scholar]
  10. Kimura K., Fujii N., Tsuzuki K., Murakami T., Indoh T., Yokosawa N., Takeshi K., Syuto B., Oguma K. 1990; The complete nucleotide sequence of the gene coding for botulinum type C1 toxin in the C-ST phage genome. Biochem Biophys Res Commun 171:1304–1311 [CrossRef]
    [Google Scholar]
  11. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685 [CrossRef]
    [Google Scholar]
  12. Maksymowych A. B., Simpson L. L. 1998; Binding and transcytosis of botulinum neurotoxin by polarized human colon carcinoma cells. J Biol Chem 273:21950–21957 [CrossRef]
    [Google Scholar]
  13. Marchesi V. T., Tillack T. W., Jackson R. L., Segrest J. P., Scott R. E. 1972; Chemical characterization and surface orientation of the major glycoprotein of the human erythrocyte membrane. Proc Natl Acad Sci USA 69:1445–1449 [CrossRef]
    [Google Scholar]
  14. Miyazaki S., Iwasaki M., Sakaguchi G. 1977; Clostridium botulinum type D toxin: purification, molecular structure, and some immunological properties. Infect Immun 17:395–401
    [Google Scholar]
  15. Moriishi K., Syuto B., Yokosawa N., Oguma K., Saito M. 1991; Purification and characterization of ADP-ribosyltransferases (exoenzyme C3) of Clostridium botulinum type C and D strains. Infect Immun 173:6025–6029
    [Google Scholar]
  16. Oguma K., Syuto B., Iida H., Kubo S. 1980; Antigenic similarity of toxins produced by Clostridium botulinum type C and D strains. Infect Immun 30:656–660
    [Google Scholar]
  17. Ohishi I., Sakaguchi G. 1980; Oral toxicities of Clostridium botulinum type C and D toxins of different molecular sizes. Infect Immun 28:303–309
    [Google Scholar]
  18. Ohishi I., Sugii S., Sakaguchi G. 1977; Oral toxicities of Clostridium botulinum toxins in response to molecular size. Infect Immun 16:107–109
    [Google Scholar]
  19. Ohyama T., Watanabe T., Fujinaga Y., Inoue K., Sunagawa H., Fujii N., Inoue K., Oguma K. 1995; Characterization of non-toxic-nonhemagglutinin component of the two types of progenitor toxin (M and L) produced by Clostridium botulinum type D CB-16. Microbiol Immunol 39:457–465 [CrossRef]
    [Google Scholar]
  20. Sakaguchi G., Ohishi I. 1984; Structure and function of botulinum toxins. In Bacterial Protein Toxins pp 435–443Edited by Alouf J. E., Fehrenbach F. J., Freer J. H., Jeljasawicz J. London: Academic Press;
    [Google Scholar]
  21. Stern M., Wentzel L. M. 1950; A new method for the large scale production of high titer botulinum formol-toxoid types C and D. J Immunol 65:175–183
    [Google Scholar]
  22. Sugii S., Ohishi I., Sakaguchi G. 1977; Correlation between oral toxicity and in vitro stability of Clostridium botulinum type A and B toxins of different molecular sizes. Infect Immun 16:910–914
    [Google Scholar]
  23. Suzuki N., Syuto B., Kubo S. 1986; Purification and characterization of hemagglutinin of Clostridium botulinum type C strain stockholm. Jpn J Vet Res 34:269–278
    [Google Scholar]
  24. Tate C. G., Tanner M. J. A. 1988; Isolation of cDNA clones for human erythrocyte membrane sialoglycoproteins α and δ. Biochem J 254:743–750
    [Google Scholar]
  25. Tsuzuki K., Kimura K., Fujii N., Yokosawa N., Indoh T., Murakami T., Oguma K. 1990; Cloning and complete nucleotide sequence of the gene for the main component of hemagglutinin produced by Clostridium botulinum type C. Infect Immun 58:3173–3177
    [Google Scholar]
  26. Tsuzuki K., Kimura K., Fujii N., Yokosawa N., Oguma K. 1992; The complete nucleotide sequence of the gene coding for the non-toxic-nonhemagglutinin component of Clostridium botulinum type C. Biochem Biophys Res Commun 183:1273–1279 [CrossRef]
    [Google Scholar]
  27. Watanabe K., Powell M. E., Hakomori S. -I. 1979; Isolation and characterization of gangliosides with a new sialosyl linkage and core structures. II. Gangliosides of human erythrocyte membranes. J Biol Chem 254:8223–8229
    [Google Scholar]
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