1887

Abstract

type C 16S progenitor toxin consists of a neurotoxin (NTX), a non-toxic non-HA (NTNH), and a haemagglutinin (HA). The HA acts as an adhesin, allowing the 16S toxin to bind to intestinal epithelial cells and erythrocytes. In type C, these bindings are dependent on sialic acid. The HA consists of four distinct subcomponents designated HA1, HA2, HA3a and HA3b. To identify the binding subcomponent(s) of HA of type C 16S toxin, all of the HA-subcomponents and some of their precursor forms were produced as recombinant proteins fused to glutathione -transferase (GST). These proteins were evaluated for their capacity to adhere to intestinal epithelial cells of guinea pig and human erythrocytes. GST-HA1, GST-HA3b and GST-HA3 (a precursor form of HA3a and HA3b) bound intestinal epithelial cells and erythrocytes, whereas GST alone, GST-HA2 and GST-HA3a did not. GST-HA3b and GST-HA3 showed neuraminidase-sensitive binding to the intestinal epithelial cells and erythrocytes, whereas GST-HA1 showed neuraminidase-insensitive binding. TLC binding assay revealed that GST-HA3b and GST-HA3 recognized sialosylparagloboside (SPG) and GM3 in the ganglioside fraction of the erythrocytes, like native type C 16S toxin [ Inoue, K. . (1999). , 2533–2542 ]. On the other hand, GST-HA1 recognized paragloboside (PG; an asialo- derivative of SPG) in addition to SPG and GM3. Deletion mutant analyses of GST-HA3b showed that the C-terminal region of HA3b is important for its binding activity. Based on these data, it is concluded that the HA component contains two distinct carbohydrate-binding subcomponents, HA1 and HA3b, which recognize carbohydrates in different specificities.

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2004-05-01
2024-03-28
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References

  1. Angata T., Brinkman-Van der Linden E. C. M. 2002; I-type lectins. Biochim Biophys Acta 1572:294–316 [CrossRef]
    [Google Scholar]
  2. Arimitsu H., Inoue K., Sakaguchi Y., Lee J., Fujinaga Y., Watanabe T., Ohyama T., Hirst R., Oguma K. 2003; Purification of fully activated Clostridium botulinum serotype B toxin for treatment of patients with dystonia. Infect Immun 71:1599–1603 [CrossRef]
    [Google Scholar]
  3. 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]
  4. Fujinaga Y., Takeshi K., Inoue K., Fujita R., Ohyama T., Moriishi K., Oguma K. 1995; Type A and B neurotoxin genes in a Clostridium botulinum type AB strain. Biochem Biophys Res Commun 213:737–745 [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 epithelial cells of guinea pig small intestine, leading to the efficient absorption of the toxin. Microbiology 143:3841–3847 [CrossRef]
    [Google Scholar]
  6. Fujinaga Y., Inoue K., Nomura T., Sasaki J., Marvard J. C., Popoff M. R., Kozaki S., Oguma K. 2000; Identification and characterization of functional subunits of Clostridium botulinum type A progenitor toxin involved in binding to intestinal microvilli and erythrocytes. FEBS Lett 467:179–183 [CrossRef]
    [Google Scholar]
  7. Hakomori S., Siddiqui B. 1974; Isolation and characterization of glycosphingolipid from animal cells and their membranes. Methods Enzymol B32:345–367
    [Google Scholar]
  8. Hauser D., Eklund M. W., Boquet P., Popoff M. R. 1994; Organisation of the botulinum neurotoxin C1 gene and its associated non-toxic protein genes in Clostridium botulinum C468. Mol Gen Genet 243:631–640
    [Google Scholar]
  9. Hazes B. 1996; The (Q X W)3 domain: a flexible lectin scaffold. Protein Sci 5:1490–1501 [CrossRef]
    [Google Scholar]
  10. Hoschützky H., Lottspeich F., Jann K. 1989; Isolation and characterization of the α-galactosyl-1,4-β-galactosyl-specific adhesin (P adhesin) from fimbriated Escherichia coli. Infect Immun 57:76–81
    [Google Scholar]
  11. 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]
  12. Inoue K., Fujinaga Y., Honke K., Yokota K., Ikeda T., Ohyama T., Takeshi K., Watanabe T., Oguma K. 1999; Characterization of haemagglutinin activity of Clostridium botulinum type C and D 16S toxins, and one subcomponent of haemagglutinin (HA1). Microbiology 145:2533–2542
    [Google Scholar]
  13. Jahn R., Niemann H. 1994; Molecular mechanisms of clostridial neurotoxins. Ann N Y Acad Sci 733:245–255 [CrossRef]
    [Google Scholar]
  14. Kouguchi H., Watanabe T., Sagane Y., Ohyama T. 2001; Characterization and reconstitution of functional hemagglutinin of the Clostridium botulinum type C progenitor toxin. Eur J Biochem 268:4019–4026 [CrossRef]
    [Google Scholar]
  15. Kouguchi H., Watanabe T., Sagane Y., Sunagawa H., Ohyama T. 2002; In vitro reconstruction of the Clostridium botulinum type D progenitor toxin. J Biol Chem 277:2650–2656 [CrossRef]
    [Google Scholar]
  16. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685 [CrossRef]
    [Google Scholar]
  17. Ledeen R. W., Yu R. K. 1982; Gangliosides: structure, isolation, and analysis. Methods Enzymol 83:139–191
    [Google Scholar]
  18. Ledeen R. W., Yu R. K., Eng L. F. 1973; Gangliosides of human myelin: sialosylgalactosylceramide (G7) as a major component. J Neurochem 21:829–839 [CrossRef]
    [Google Scholar]
  19. Magnani J. L., Nilsson B., Brockhaus M., Zopf D., Steplewaki Z., Koprowski H., Ginsburg V. 1982; A monoclonal antibody-defined antigen associated with gastrointestinal cancer is a ganglioside containing sialylated lacto-N-fucopentaose II. J Biol Chem 257:14365–14369
    [Google Scholar]
  20. 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]
  21. May A. P., Robinson R. C., Vinson M., Crocker P. R., Jones E. Y. 1998; Crystal structure of the N-terminal domain of sialoadhesin in complex with 3′ sialyllactose at 1·85 Å resolution. Mol Cell 1:719–728 [CrossRef]
    [Google Scholar]
  22. Minton N. P. 1995; Molecular genetics of clostridial neurotoxins. Curr Top Microbiol Immunol 95:161–194
    [Google Scholar]
  23. Montecucco C., Schiavo G. 1994; Mechanism of action of tetanus and botulism neurotoxins. Mol Microbiol 13:1–8 [CrossRef]
    [Google Scholar]
  24. Mahmut N., Inoue K., Fujinaga Y. & 7 other authors; 2002; Characterisation of monoclonal antibodies against haemagglutinin associated with Clostridium botulinum type C neurotoxin. J Med Microbiol 51:286–294
    [Google Scholar]
  25. 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]
  26. Oguma K., Fujinaga Y., Inoue K. 1995; Structure and function of Clostridium botulinum toxins. Microbiol Immunol 39:161–168 [CrossRef]
    [Google Scholar]
  27. Oguma K., Fujinaga Y., Inoue K. 1997; Clostridium botulinum toxin. J Toxicol Toxin Rev 16:253–266 [CrossRef]
    [Google Scholar]
  28. Oguma K., Inoue K., Fujinaga Y., Yokota K., Watanabe T., Ohyama T., Takeshi K., Inoue K. 1999; Structure and function of Clostridium botulinum progenitor toxin. J Toxicol Toxin Rev 18:17–34 [CrossRef]
    [Google Scholar]
  29. Ohyama T., Watanabe T., Fujinaga Y., Inoue K., Sunagawa H., Fujii N., Inoue K., Oguma K. 1995; Characterization of nontoxic-nonhemaggulutinin component of two types of progenitor toxin (M and L) produced by Clostridium botulinum type D CB-16. Microbiol Immunol 39:457–465 [CrossRef]
    [Google Scholar]
  30. Sagane Y., Kouguchi H., Watanabe T., Sunagawa H., Inoue K., Fujinaga Y., Oguma K., Ohyama T. 2001; Role of C-terminal region of HA-33 component of Botulinum toxin in hemagglutination. Biochem Biophys Res Commun 288:650–657 [CrossRef]
    [Google Scholar]
  31. Sakaguchi G., Kozaki S., Ohishi I. 1984; Structure and function of botulinum toxins. In Bacterial Protein Toxins pp. 435–443 London: Academic Press;
    [Google Scholar]
  32. Schiavo G., Matteoli M., Montecucco C. 2000; Neurotoxin affecting neuroexocytosis. Physiol Rev 80:717–766
    [Google Scholar]
  33. 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 hemaggulutinin produced by Clostridium botulinum type C. Infect Immun 58:3173–3177
    [Google Scholar]
  34. Uemura K., Yazawa M., Taketomi T. 1978; Characterization of major glycolipids in bovine erythrocyte membrane. J Biochem 83:463–471
    [Google Scholar]
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