Glycoconjugate Journal

, Volume 16, Issue 7, pp 327–336 | Cite as

UDP-N-Acetyl-α-D-glucosamine as acceptor substrate of β-1,4-galactosyltransferase. Enzymatic synthesis of UDP-N-acetyllactosamine

  • Lothar Elling
  • Astrid Zervosen
  • Ricardo Gutiérrez Gallego
  • Veronika Nieder
  • Martine Malissard
  • Eric G. Berger
  • Johannes F.G. Vliegenthart
  • Johannis P. Kamerling

Abstract

The capacity of UDP-N-acetyl-α-D-glucosamine (UDP-GlcNAc) as an in vitro acceptor substrate for β-1,4-galactosyltransferase (β4GalT1, EC 2.4.1.38) from human and bovine milk and for recombinant human β4GalT1, expressed in Saccharomyces cerevisiae, was evaluated. It turned out that each of the enzymes is capable to transfer Gal from UDP-α-D-galactose (UDP-Gal) to UDP-GlcNAc, affording Gal(β1-4)GlcNAc(α1-UDP (UDP-LacNAc). Using β4GalT1 from human milk, a preparative enzymatic synthesis of UDP-LacNAc was carried out, and the product was characterized by fast-atom bombardment mass spectrometry and 1H and 13C NMR spectroscopy. Studies with all three β4GalTs in the presence of α-lactalbumin showed that the UDP-LacNAc synthesis is inhibited and that UDP-α-D-glucose is not an acceptor substrate. This is the first reported synthesis of a nucleotide-activated disaccharide, employing a Leloir glycosyltransferase with a nucleotide-activated monosaccharide as acceptor substrate. Interestingly, in these studies β4GalT1 accepts an α-glycosidated GlcNAc derivative. The results imply that β4GalT1 may be responsible for the biosynthesis of UDP-LacNAc, previously isolated from human milk.

glycosyltransferase β-1,4-galactosyltransferase nucleotide-activated disaccharide UDP-LacNAc BSA, bovine serum albumin FAB-MS, Fast Atom Bombardment Mass Spectrometry 2D HMBC, two-dimensional Heteronuclear Multiple-Bond Coherence 2D ROESY, two-dimensional Rotating Frame Nuclear Overhauser Enhancement Spectroscopy 2D TOCSY, two-dimensional Total Correlation Spectroscopy β4GalT, β-1,4-galactosyltransferase α-LA, α-lactalbumin LacNAc, N-acetyllactosamine NMR, Nuclear Magnetic Resonance UDP-GlcNAc, uridine 5′-diphospho-N-acetyl-α-D-glucosamine UDP-Gal, uridine 5′-diphospho-α-D-galactose UDP-GalNAc, uridine 5′-diphospho-N-acetyl-α-D-galactosamine UDP-Glc, uridine 5′-diphospho-α-D-glucose UDP-LacNAc, uridine 5′-diphospho-N-acetyllactosamine UDP-Xyl, uridine 5′-diphospho-α-D-xylose 

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References

  1. 1.
    Trayer IP, Hill RL (1971) J Biol Chem 246: 6666–75.Google Scholar
  2. 2.
    Andrews P (1970) FEBS Lett 9: 297–300.Google Scholar
  3. 3.
    Gerber AC, Kozdrowski I, Wyss S, Berger EG (1979) Eur J Biochem 93: 453–60.Google Scholar
  4. 4.
    Brodbeck U, Ebner KE (1966) J Biol Chem 241: 762–64.Google Scholar
  5. 5.
    Ebner KE (1971) J Dairy Sci 54: 1229–33.Google Scholar
  6. 6.
    Hill RL, Brew K (1975) Adv Enzymol 43: 411–90.Google Scholar
  7. 7.
    Almeida R, Amado M, David L, Levery SB, Holmes EH, Merkx G, van Kessel AG, Rygaard E, Hassan H, Bennett E, Clausen H (1997) J Biol Chem 272: 31979–91.Google Scholar
  8. 8.
    Wong C-H, Ichikawa Y, Krach T, Gautheron-Le Narvor C, Dumas DP, Look GC (1991) J Am Chem Soc 113: 8137–45.Google Scholar
  9. 9.
    David S, Augé C, Gautheron C (1991) Adv Carbohydr Chem Biochem 49: 175–237.Google Scholar
  10. 10.
    Yoon E, Laine RA (1992) Glycobiology 2: 161–8.Google Scholar
  11. 11.
    Wong C-H, Whitesides GM (1994) Enzymes in Synthetic Organic Chemistry. Oxford: Elsevier Science.Google Scholar
  12. 12.
    Palcic MM, Hindsgaul O (1996) Trends Glycosci Glycotechnol 8: 37–49.Google Scholar
  13. 13.
    Baisch G, Öhrlein R, Ernst B (1996) Bioorg Med Chem Lett 6: 749–54.Google Scholar
  14. 14.
    Gambert U, Thiem J (1997) Topics Curr Chem 186: 21–43.Google Scholar
  15. 15.
    Zervosen A, Elling L (1996) J Am Chem Soc 118: 1836–40.Google Scholar
  16. 16.
    Hokke CH, Zervosen A, Elling L, Joziasse DH, van den Eijnden DH (1996) Glycoconjugate J 13: 687–92.Google Scholar
  17. 17.
    Kobata A (1963) J Biochem 53: 167–75.Google Scholar
  18. 18.
    Jourdian GW, Shimizu F, Roseman S (1961) Fed Proc 20: 161.Google Scholar
  19. 19.
    Suzuki S (1962) J Biol Chem 237: 1393.Google Scholar
  20. 20.
    Nakanishi Y, Shimizu S, Takahashi N, Sugiyama M, Suzuki S (1967) J Biol Chem 242: 967–76.Google Scholar
  21. 21.
    Hartmann E, König H (1989) Arch Microbiol 151: 274–81.Google Scholar
  22. 22.
    Hartmann E, König H (1991) Biol Chem Hoppe Seyler 372: 971–74.Google Scholar
  23. 23.
    Hartmann E, Messner P, Allmeier G, König H (1993) J Bacteriol 175: 4515–19.Google Scholar
  24. 24.
    König H, Kandler O, Hammes W (1989) Can J Microbiol 35: 176–81.Google Scholar
  25. 25.
    König H, Hartmann E, Kärcher U (1994) Syst Appl Microbiol 16: 510–17.Google Scholar
  26. 26.
    Bülter T, Wandrey C, Elling L (1997) Carbohydr Res 305: 469–73.Google Scholar
  27. 27.
    Malissard M, Borsig L, Di Marco S, Grütter MG, Kragl U, Wandrey C, Berger EG (1996) Eur J Biochem 239: 340–48.Google Scholar
  28. 28.
    Elling L, Kula M-R (1993) J Biotechnol 29: 277–86.Google Scholar
  29. 29.
    Hård K, van Zadelhoff G, Moonen P, Kamerling JP, Vliegenthart JFG (1992) Eur J Biochem 209: 895–915.Google Scholar
  30. 30.
    Summers MF, Marzilli LG, Bax A (1986) J Am Chem Soc 108: 4285–94.Google Scholar
  31. 31.
    Unverzagt C, Paulson JC (1990) J Am Chem Soc 112: 9308–09.Google Scholar
  32. 32.
    Wong C-H, Krach T, Gautheron-Le Narvor C, Ichikawa Y, Look GC, Gaeta F, Thompson D, Nicolaou KC (1991) Tetrahedr Lett 32: 4867–70.Google Scholar
  33. 33.
    Takayama S, Shimazaki M, Qiao L, Wong C-H (1996) Bioorg Med Chem Lett 6: 1123–26.Google Scholar
  34. 34.
    Yoon JH, Ajisaka K (1996) Carbohydr Res 292: 153–63.Google Scholar
  35. 35.
    Köplin R, Brisson J-R, Whitfield C (1997) J Biol Chem 272: 4121–28.Google Scholar
  36. 36.
    Khatra BS, Herries DG, Brew K (1974) Eur J Biochem 44: 537–60.Google Scholar
  37. 37.
    Bell JE, Beyer TA, Hill RL (1976) J Biol Chem 251: 3003–13.Google Scholar
  38. 38.
    Morrison JF, Ebner KE (1971) J Biol Chem 246: 3977–84.Google Scholar
  39. 39.
    Nunez HA, Barker R (1976) Biochemistry 15: 3843–47.Google Scholar
  40. 40.
    Brew K, Vanaman TC, Hill RL (1968) Proc Natl Acad Sci USA 59: 491–97.Google Scholar
  41. 41.
    Klee WA, Klee CB (1970) Biochem Biophys Res Commun 39: 833–41.Google Scholar
  42. 42.
    Berliner LJ, Davis ME, Ebner KE, Beyer TA, Bell JE (1984) Mol Cell Biochem 62: 37–42.Google Scholar
  43. 43.
    Babad H, Hassid WZ (1966) J Biol Chem 241: 2672–78.Google Scholar
  44. 44.
    Do K-Y, Do S-I, Cummings RD (1995) J Biol Chem 270: 18447–51.Google Scholar
  45. 45.
    Andree PJ, Berliner LJ (1978) Biochim Biophys Acta 544: 489–95.Google Scholar
  46. 46.
    Kren V, Augé C, Sedmera P, Havlicek V (1994) Chem Soc Perkin Trans 1 : 2481–84.Google Scholar
  47. 47.
    Crawley SC, Palcic MM (1996) In Modern Methods in Carbohydrate Synthesis (Khan SH, O'Neill RA, eds) pp 492–517. Amsterdam: Harwood Academic Publishers.Google Scholar
  48. 48.
    Palcic MM, Hindsgaul O (1991) Glycobiology 1: 205–09.Google Scholar
  49. 49.
    Schwientek T, Almeida R, Levery SB, Holmes EH, Bennett E, Clausen H (1998) Biol Chem 273: 29331–40.Google Scholar
  50. 50.
    Sato T, Furukawa K, Bakker H, van den Eijnden DH, van Die I (1998) Proc Natl Acad Sci USA 95: 472–77.Google Scholar
  51. 51.
    Sato T, Aoki N, Matsuda T, Furukawa K (1998) Biochem Biophys Res Commun 244: 637–41.Google Scholar
  52. 52.
    Nomura T, Takizawa M, Aoki J, Arai H, Inoue K, Wakisaka E, Yoshizuka N, Imokawa G, Dohmae N, Takio K, Hattori M, Matsuo N (1998) J Biol Chem 273: 13570–77.Google Scholar
  53. 53.
    Ujita M, McAuliffe J, Schwientek T, Almeida R, Hindsgaul O, Clausen H, Fukuda M (1998) J Biol Chem 273: 34843–49.Google Scholar

Copyright information

© Kluwer Academic Publishers 1999

Authors and Affiliations

  • Lothar Elling
    • 1
  • Astrid Zervosen
    • 1
  • Ricardo Gutiérrez Gallego
    • 2
  • Veronika Nieder
    • 1
  • Martine Malissard
    • 3
  • Eric G. Berger
    • 3
  • Johannes F.G. Vliegenthart
    • 2
  • Johannis P. Kamerling
    • 2
  1. 1.Institute of Enzyme TechnologyHeinrich-Heine-University Düsseldorf, Research Center JülichJülichGermany
  2. 2.Bijvoet Center, Department of Bio-Organic ChemistryUtrecht UniversityUtrechtThe Netherlands
  3. 3.Institute of PhysiologyUniversity of ZürichZürichSwitzerland

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