Biosynthesis of Glycophorin A

  • Odd Nygård
  • Peter Westermann

Abstract

The major human red cell sialoglycoprotein, glycophorin A, is structurally one of the best known membrane proteins. Until recently, however, nothing was known about its biosynthesis. We found in 1979 that the K562 cell line is erythroid and synthesizes glycophorin A. Glycophorin A mRNA isolated from K562 cells directed in vitro the synthesis of a protein with an apparent molecular weight of 19500. That exceeded the molecular weight of the apoprotein with 5000, indicating the presence of a signal peptide. Glycophorin A synthesized in vitro in the presence of dog pancreatic membranes had an apparent molecular weight of 37000. This precursor bound to lentil lectin but not to Helix pomatia lectin. Pulse-chase experiments in vivo showed that initially a 37000 molecular weight protein, which bound to lentil lectin, was synthesized which gradually was replaced by a 39000 molecular weight protein. Glycosylation was completed in about 10 min and the protein appeared at the cell surface after 25 min. Inhibition of glycosylation of the single K-glycosidic oligosaccharide ty tunicamycin did not affect the migration of the protein to the cell surface. Using the N-acetylgalactosamine specific lectin from Helix pomatia coupled to Sepharose two early N-acety]galactosamine-containing O-giycosylated glycophorin A precursors with apparent molecular weights of 24000 and 27000 were identified. O-glycosylation took place in the presence of carboxylic ionophore monensin, but the protein stained intracellularly. The relationship between the 37000 and 24000–27000 molecular weight proteins remains unclear but the results indicate that the synthesis of O-glycosylated glycoproteins is more complex than generally believed and may involve early uncharacterized precursors.

Keywords

Serine Polypeptide Trypsin Methionine Oligosaccharide 

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References

  1. Andersson, L.C., Jokinen, M. & Gahmberg, C.G. (1979) Nature 278, 364–365.PubMedCrossRefGoogle Scholar
  2. Blobel, G. (1960) Proc. Natl. Acad. Sci. USA 77, 1496–1500.CrossRefGoogle Scholar
  3. Dahr, W., Uhlenbruck, G., Leikola, J., Wagstaff, W. & Landfried, K. (1976) Immunogenet. 3, 329–346.CrossRefGoogle Scholar
  4. Gahmberg, C.G. & Andersson, L.C. (1977) J. Biol. Chem. 252, 5888–5894.PubMedGoogle Scholar
  5. Gahmberg, C.G. & Andersson, L.C. (1982) Eur. J. Biochem. 122, 581–586.PubMedCrossRefGoogle Scholar
  6. Gahmberg, C.G., Jokinen, M. & Andersson, L.C. (1978) Blood 52, 379–387.PubMedGoogle Scholar
  7. Gahmberg, C.G., Jokinen, M. & Andersson, L.C. (1979) J. Biol. Chem. 254, 7442–7448.PubMedGoogle Scholar
  8. Gahmberg, C.G., Jokinen, M., Karhi, K.K. & Andersson, L.C. (1980) J. Biol. Chem. 255, 2169–2175.PubMedGoogle Scholar
  9. Gahmberg, C.G., Jokinen, M., Karhi, K.K., Kämpe, O., Peterson, P.A. & Andersson, L.C. (1980) Methods Enzymol. in press.Google Scholar
  10. Gahmberg, C.G., Jokinen, M., Karhi, K.K., Ulmanen, I., Kääriäinen, L. & Andersson, L.C. (1981) Blood Transfusion and Immunohaematology XXIV, 53–73.Google Scholar
  11. Gahmberg, C.G., Myllylä, G., Leikola, J., Pirkola, A. & Nordling, S. (1976) J. Biol. Chem. 251, 6108–6116.PubMedGoogle Scholar
  12. Hanover, J.A., Lennarz, W.J. & Young, J.D. (1980) J. Biol. Chem. 255, 6713–6716.PubMedGoogle Scholar
  13. Hubbard, S.V., & Ivatt, R.J., (1981) Annu. Rev. Biochem. 50, 555–583.PubMedCrossRefGoogle Scholar
  14. Johnson, D.C. & Spear, P.G. (1983) Cell 32, 987–997.PubMedCrossRefGoogle Scholar
  15. Jokinen, M., Gahrnberg, C.G. & Andersson, L.C. (1979) Nature 279, 604–607.PubMedCrossRefGoogle Scholar
  16. Jokinen, M., Ulmanen, I., Andersson, L.C., Kääriäinen, L. & Gahrnberg, C.G. (1981) Eur. J. Biochem. 114, 393–397.PubMedCrossRefGoogle Scholar
  17. Lodish, H.F. & Rothman, J.E. (1979) Sci. American 240, 48–63.CrossRefGoogle Scholar
  18. Meyer, D.I., Krause, E. & Dobberstein, B. (1982) Nature 297, 647–650.PubMedCrossRefGoogle Scholar
  19. Nieman, H., Boschek, B., Evans, D., Rosing, M., Tamura, T. & Klenk, H.-D. (1983) EMB0 Journal 1, 1499–1504.Google Scholar
  20. Rutherford, T.R., Clegg, J.B. & Wheaterall, D.J. (1979) Nature 280, 164–165.PubMedCrossRefGoogle Scholar
  21. Tabas, I. & Kornfeld, S. (1979) J. Biol. Chem. 254, 11655– 11663.PubMedGoogle Scholar
  22. Tanner, M.J.A. & Anstee, D.J. (1976) Biochem. J. 153, 271– 277.PubMedGoogle Scholar
  23. Tartakoff, A.M. (1983) Cell 32, 1026–1028.PubMedCrossRefGoogle Scholar
  24. Tulsiani, D.R.P., Hubbard, S.C., Robbins, P.W. & Touster, O. (1982) J. Biol. Chem. 257, 3660–3668.PubMedGoogle Scholar
  25. Villa-Komaroff, L., McDowell, M., Baltimore, D. & Lodish, H.L. (1974) Methods Enzymol. 30, 709–723.PubMedCrossRefGoogle Scholar
  26. Walter, P. & Blobel, G. (1982) Nature 299, 691–698.PubMedCrossRefGoogle Scholar

Copyright information

© The Human Press Inc. 1983

Authors and Affiliations

  • Odd Nygård
    • 1
  • Peter Westermann
    • 2
  1. 1.The Wenner-Gren InstituteUniversity of StockholmStockholmSweden
  2. 2.Central Institute of Molecular BiologyAcademy of Sciences of GDRBerlin-BuchGermany

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