1887

Abstract

Uridine-5′-diphospho--acetylgalactosamine (UDP-GalNAc) is required in the formation of the outer filamentous wall of and is synthesized by inducible enzymes in the cytosol of encysting trophozoites. In this study, an inducible enzyme activity that is associated with a particle population isolated from encysting is reported, and this activity exclusively incorporates [1-C]GalNAc (from UDP-[C]GalNAc) into an ethanol precipitate with the same properties as the filamentous cyst wall of . This ethanol precipitate exhibits characteristics of cyst wall filaments in that both contain GalNAc as the only sugar moieties and are SDS-insoluble, proteinase- and alkali-resistant and acid-hydrolysable. However, since the precise chemical nature of the ethanol precipitate remains unknown, this enzyme activity is referred to tentatively as cyst wall synthase (CWS). CWS activity peaks in cells between 24 and 36 h of encystment and exhibits a high affinity and marked specificity for UDP-GalNAc as its substrate. UDP--acetylglucosamine, UDP-glucose, UDP-galactose, -glucosamine and -galactosamine were not incorporated into the ethanol precipitate. Partially purified CWS activity exhibits an apparent of 0·048 mM for UDP-GalNAc, a of 0·70 nmol min (mg protein) and a requirement for divalent cations in the following order of preference: Ca, Mg>Co>>>Mn, Zn. EDTA inhibits CWS activity.

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2004-05-01
2019-11-21
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References

  1. Boone, J. H., Wilkins, T. D., Nash, T. E., Brandon, J. E., Macias, E. A., Jerris, R. C. & Lyerly, D. M. ( 1999; ). TechLab and Alexon Giardia enzyme-linked immunosorbent assay kits detect cyst wall protein 1. J Clin Microbiol 37, 611–614.
    [Google Scholar]
  2. Das, S. & Gillin, F. D. ( 1996; ). Giardia lamblia: increased UDP-N-acetyl-d-glucosamine and udp-n-acetyl-d-galactosamine transferase activities during encystation. Exp Parasitol 83, 19–29.[CrossRef]
    [Google Scholar]
  3. DeAngelis, P. L. ( 1999; ). Hyaluronan synthases: fascinating glycosyltransferases from vertebrates, bacterial pathogens, and algal viruses. Cell Mol Life Sci 56, 670–682.[CrossRef]
    [Google Scholar]
  4. Duran, A. & Cabib, E. ( 1978; ). Solubilization and partial purification of yeast chitin synthase. Confirmation of the zymogenic nature of the enzyme. J Biol Chem 253, 4419–4425.
    [Google Scholar]
  5. Erlandsen, S. L., Bemrick, W. J. & Pauley, J. ( 1989; ). High-resolution electron microscopic evidence for the filamentous structure of the cyst wall in Giardia muris and Giardia duodenalis. J Parasitol 75, 787–797.[CrossRef]
    [Google Scholar]
  6. Faubert, G., Reiner, D. S. & Gillin, F. D. ( 1991; ). Giardia lamblia: regulation of secretary vesicle formation and loss of ability to reattach during encystation in vitro. Exp Parasitol 72, 345–354.[CrossRef]
    [Google Scholar]
  7. Feely, D. E., Erlandsen, S. L. & Chase, D. G. ( 1984; ). Structure of the trophozoite and cyst. In Giardia and Giardiasis, pp. 3–31. Edited by S. Erlandsen & E. Meyer. New York: Plenum.
  8. Gerwig, G. J., van Kuik, J. A., Leeflang, B. R., Kamerling, J. P., Vliegenthart, J. F., Karr, C. D. & Jarroll, E. L. ( 2002; ). The Giardia intestinalis filamentous cyst wall contains a novel β(1-3)-N-acetyl-d-galactosamine polymer: a structural and conformational study. Glycobiology 12, 499–505.[CrossRef]
    [Google Scholar]
  9. Hare, D. F., Jarroll, E. L. & Lindmark, D. G. ( 1989; ). Giardia lamblia: characterization of proteinase activity in trophozoites. Exp Parasitol 68, 168–175.[CrossRef]
    [Google Scholar]
  10. Jarroll, E. L., Manning, P., Lindmark, D. G., Coggins, J. R. & Erlandsen, S. L. ( 1989; ). Giardia cyst wall-specific carbohydrate: evidence for the presence of galactosamine. Mol Biochem Parasitol 32, 121–132.[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. Lane, S. & Lloyd, D. ( 2002; ). Current trends in research into the waterborne parasite Giardia. Crit Rev Microbiol 28, 123–147.[CrossRef]
    [Google Scholar]
  13. Lindmark, D. G. ( 1988; ). Giardia lamblia: localization of hydrolase activities in lysosome-like organelles of trophozoites. Exp Parasitol 65, 141–147.[CrossRef]
    [Google Scholar]
  14. Lujan, H. D., Marotta, A., Mowatt, M. R., Sciaky, N., Lippincott-Scwartz, J. & Nash, T. E. ( 1995a; ). Developmental induction of Golgi structure and function in the primitive eukaryote Giardia lamblia. J Biol Chem 270, 4612–4618.[CrossRef]
    [Google Scholar]
  15. Lujan, H. D., Mowatt, M. R., Conrad, J. T., Bowers, B. & Nash, T. E. ( 1995b; ). Identification of a novel Giardia lamblia cyst wall protein with leucine-rich repeats. Implications for secretory granule formation and protein assembly into the cyst wall. J Biol Chem 270, 29307–29313.[CrossRef]
    [Google Scholar]
  16. Macechko, P. T., Steimle, P. A., Lindmark, D. G., Erlandsen, S. L. & Jarroll, E. L. ( 1992; ). Galactosamine-synthesizing enzymes are induced when Giardia encyst. Mol Biochem Parasitol 56, 301–310.[CrossRef]
    [Google Scholar]
  17. Manning, P., Erlandsen, S. L. & Jarroll, E. L. ( 1992; ). Carbohydrate and amino acid analyses of Giardia muris cysts. J Protozool 39, 290–296.[CrossRef]
    [Google Scholar]
  18. Mowatt, M. R., Lujan, H. D., Cotten, D. B., Bowers, B., Yee, J., Nash, T. E. & Stibbs, H. H. ( 1995; ). Developmentally regulated expression of a Giardia lamblia cyst wall protein gene. Mol Microbiol 15, 955–963.[CrossRef]
    [Google Scholar]
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