New Techniques in Glycosyltransferase Research

  • Manju Basu
  • Kamal K. Das
  • Hung-Che Chon
  • Subhash Basu
Part of the NATO ASI Series book series (NSSA, volume 116)


Glycosyltransferases (GLTs) of eukaryotic origin catalyze the transfer of glycose units to the appropriate glycoprotein, ceramide, or glycolipid core structures. These enzymes may recognize a specific terminal glycose unit or the penultimate sugar, in addition to the terminal sugar unit.1–4 GLTs are believed to be localized in the Golgi bodies,5–7 and most of them are membrane-bound. Many have been solubilized using various detergents, and their purification is under way.8–12 We are involved in the purification of glycolipid:glycosyltransferases (GSL:GLTs) and in the study of their kinetic properties.8–13 This article describes relatively easy methods for isolation14,15 of glycolipid substrates for GSL:GLTs and the development of some convenient, relatively inexpensive assay methods.16 Both advances were essential before substantial numbers of fractions obtained during column chromatography could be assayed.


Embryonic Chicken Sodium Metabisulfite Rabbit Bone Marrow Palmitoyl Chloride Buffy Coat Layer 
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.
    S. Roseman, The synthesis of complex carbohydrates by multiglycosyltransferase systems and their potential function in intercellular adhesion, Chem. Phys. Lipid, 5, 270 (1970).CrossRefGoogle Scholar
  2. 2.
    S. Basu, M. Basu, J.W..Kyle, and H.C. Chon, Biosynthesis in vitro of gangliosides containing Gg-and Lc-cores, in “Ganglioside Structure and Function” by (eds. R. Ledeen, M. Rapport, K. Suzuki, and R. Yu) Plenum Press, 249 (1984).Google Scholar
  3. 3.
    S. Basu and M. Basu, Expression of glycosphingolipid glycosyltransferases in development and transformation, in “The Glycoconjugates” by (ed. M. Horowitz) Academic Press, New York, Vol. 3, 265 (1982).Google Scholar
  4. 4.
    T.A. Beyer and R.L. Hill, Glycosylation pathways in the biosynthesis of nonreducing terminal sequences in oligosaccharides of glycoproteins, in “The Glycoconjugates” by (ed. M. Horowitz) Academic Press, New York, Voo. 3, 25 (1982).Google Scholar
  5. 5.
    R.L. Hudgin, R.K. Murray, L. Pinteric, H.P. Morris and H. Schachter, The use of nucleotide sugar: glycoprotein glycosyltransferases to asses Golgi apparatus function in Morris Hepatomas, Canad. J. Biochem., 49, 61 (1971).Google Scholar
  6. 6.
    T.W. Keenan, D.J. Morre, and S. Basu, Ganglioside biosynthesis: concentration of glycosphingolipid glycosyltransferases in Golgi apparatus from rat liver, J. Biol. Chem., 249, 310 (1974).PubMedGoogle Scholar
  7. 7.
    E.G. Berger, T. Mande, and U. Schilt, Immunohistochemical localization of galactosyltransferase in human fibroblasts and HeLa cells, J. Histo. Cytochem., 29, 364 (1981).CrossRefGoogle Scholar
  8. 8.
    S. Basu, M. Basu, J.W. Kyle, T. De, K. Das, and R.J. Schaeper, in “Enzymes of Lipid Metabolism” by (eds. L. Freysz and S. Gatt) (in press) Plenum Press, New York.Google Scholar
  9. 9.
    S. Basu, T. De, J.W. Kyle, and M. Basu, Biosynthesis of eukaryotic cell surface glycosphingolipids using solubilized glycosyltransferases, Proc. Int. Symp. Biomol. Struc. Interactions, Suppl. J. Biosci., 8 (in press).Google Scholar
  10. 10.
    J.W. Kyle, Characterization of glycolipid galactosyltransferases from embryonic chicken brain, Ph.D. Thesis, University of Notre Dame, Notre Dame, IN (1985).Google Scholar
  11. 11.
    H. Higashi, M. Basu, and S. Basu, Biosynthesis in vitro of disialoneolactotetraosylceramide by a solubilized sialyltransferase from embryonic chicken brain, J. Biol. Chem., 260, 824 (1985).PubMedGoogle Scholar
  12. 12.
    M. Basu and S. Basu, Biosynthesis in vitro of Ii-core glycolipids from neolactotetraosylceramide by 81–3 and 81–6 N-acetylglucosaminyltransferases from mouse T-lymphoma, J. Biol. Chem., 259, 12557 (1984).PubMedGoogle Scholar
  13. 13.
    H.C. Chon, Solubilization of glycolipid sialyltransferases from embryonic chicken liver and brain. Use of radiolabeled glycolipids in vitro to assay sialyltransferases, M.S. Thesis, University of Notre Dame, Notre Dame, IN (1984).Google Scholar
  14. 14.
    M. Basu and S. Basu, Enzymatic synthesis of blood group B specific pentaglycosylceramide by an a-galactosyltransferase from rabbit bone marrow, J. Biol. Chem., 257, 169 (1983).Google Scholar
  15. 15.
    K.K. Das, M. Basu, and S. Basu, A rapid preparative method for isolation of neutral and acidic glycosphingolipids by radial thin-layer chromatography, Anal. Biochem., 143, 125 (1984).PubMedCrossRefGoogle Scholar
  16. 16.
    M. Basu, T. De, K.K. Das, J.W. Kyle, H.C. Chon, R.J. Schaeper, and S. Basu, Glycosyltransferases involved in glycolipid biosynthesis, Methods in Enzymol (in press).Google Scholar
  17. 17.
    M. Basu, S. Basu, A. Stoffyn, and P. Stoffyn, Biosynthesis in vitro of sialyl(a2–3)neolactotetraosylceramide by a sialyltransferase from embryonic chicken brain, J. Biol. Chem., 257, 12765 (1982).PubMedGoogle Scholar
  18. 18.
    H. Higashi and S. Basu, Specific 14C-labeling of sialic acid and Nacetylhexosamine residues of glycosphingolipids after hydrazinolysis, Anal. Biochem., 120, 159 (1982).PubMedCrossRefGoogle Scholar
  19. 19.
    Y. Suzuki, Y. Hirabayashi, and M. Matsumoto, Hydrazinolysis of glycosphingolipids. A new method for preparation of N-deacylated (lyso) glycosphingolipids, J. Biochem., 95, 1219 (1984).PubMedGoogle Scholar
  20. 20.
    J.E. Sadler, T.A. Beyer, C.L. Oppenheimer, J.C. Paulson, J.L. Prieels, J.I. Rearick, and R.L. Hill, Purification of mammalian glycosyltransferases, Methods Enzymol., 81, 458 (1982).CrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1986

Authors and Affiliations

  • Manju Basu
    • 1
  • Kamal K. Das
    • 1
  • Hung-Che Chon
    • 1
  • Subhash Basu
    • 1
  1. 1.Department of Chemistry Biochemistry, Biophysics and Molecular Biology ProgramUniversity of Notre DameNotre DameUSA

Personalised recommendations