Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1979 Jun 15;180(3):481–489. doi: 10.1042/bj1800481

The effect of tunicamycin on the glycosylation of lactating-rabbit mammary glycoproteins

Brian K Speake 1, David A White 1
PMCID: PMC1161085  PMID: 486126

Abstract

1. Tunicamycin inhibited the incorporation of d-[2-3H]mannose into dolichol-linked oligosaccharide and glycoprotein of lactating-rabbit mammary explants by approximately the same extent (approx. 30% of control value), suggesting that lipid-linked intermediates are involved in the mannosylation of mammary glycoproteins. 2. The incorporation of radioactivity from N-acetyl-d-[1-14C]glucosamine into dolichol-linked oligosaccharide was inhibited by tunicamycin to 32% of the control value, whereas the incorporation of the radiolabel into glycoprotein was only inhibited to 72% of the control value. 3. Considerable redistribution of label from N-acetylglucosamine to N-acetylgalactosamine was found to occur in the explants. In the presence of tunicamycin approx. 76% of the radioactivity incorporated into glycoprotein from N-acetyl-d-[1-14C]glucosamine was present as N-acetylgalactosamine, compared with approx. 61% in the absence of the inhibitor. Thus tunicamycin selectively inhibits the incorporation of N-acetylglucosamine into glycoprotein. 4. Radioactivity from N-acetyl-d-[1-14C]glucosamine was incorporated into a glycoprotein that was identified as casein by the use of a casein-specific antiserum, and also into a group of glycopolypeptides with apparent mol.wts. ranging between 40000 and 80000. N-Acetylgalactosamine was the only radioactive sugar released on strong-acid hydrolysis of the immunoprecipitated casein, whereas N-acetylglucosamine was the major radioactive residue present in the non-casein glycoproteins. Glucosamine and galactosamine were the only radiolabelled sugars detected by paper chromatography of the strong-acid hydrolysate of the protein fraction. 5. Tunicamycin inhibited the incorporation of radioactivity from N-acetyl-d-[1-14C]glucosamine into the glycopolypeptides with mol.wts. between 40000 and 80000 as described by polyacrylamide-gel electrophoresis, but did not affect the incorporation of label into casein. It appears that tunicamycin inhibits the incorporation of mannose and N-acetylglucosamine into a number of mammary glycoproteins by inhibiting the formation of lipid-linked intermediates, but does not inhibit the incorporation of N-acetylgalactosamine into casein.

Full text

PDF
481

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Al-Sarraj K., White D. A., Mayer R. J. Immunochemical characterization of casein from rabbit mammary gland. Biochem J. 1978 Sep 1;173(3):877–883. doi: 10.1042/bj1730877. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Al-Sarraj K., White D. A., Mayer R. J. Purification and properties of casein from mammary gland of lactating rabbits. Int J Biochem. 1978;9(4):269–277. doi: 10.1016/0020-711x(78)90009-5. [DOI] [PubMed] [Google Scholar]
  3. Duksin D., Bornstein P. Changes in surface properties of normal and transformed cells caused by tunicamycin, an inhibitor of protein glycosylation. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3433–3437. doi: 10.1073/pnas.74.8.3433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Eagon P. K., Heath E. C. Glycoprotein biosynthesis in myeloma cells. Characterization on nonglycosylated immunoglobulin light chain secreted in presence of 2-deoxy-D-glucose. J Biol Chem. 1977 Apr 10;252(7):2372–2383. [PubMed] [Google Scholar]
  5. Hemming F. W. Dolichol phosphate, a coenzyme in the glycosylation of animal membrane-bound glycoproteins. Biochem Soc Trans. 1977;5(4):1223–1231. doi: 10.1042/bst0051223. [DOI] [PubMed] [Google Scholar]
  6. Hickman S., Kulczycki A., Jr, Lynch R. G., Kornfeld S. Studies of the mechanism of tunicamycin in hibition of IgA and IgE secretion by plasma cells. J Biol Chem. 1977 Jun 25;252(12):4402–4408. [PubMed] [Google Scholar]
  7. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  8. Lehle L., Tanner W. The specific site of tunicamycin inhibition in the formation of dolichol-bound N-acetylglucosamine derivatives. FEBS Lett. 1976 Nov 15;72(1):167–170. doi: 10.1016/0014-5793(76)80922-2. [DOI] [PubMed] [Google Scholar]
  9. MORGAN J. F., MORTON H. J., PARKER R. C. Nutrition of animal cells in tissue culture; initial studies on a synthetic medium. Proc Soc Exp Biol Med. 1950 Jan;73(1):1–8. doi: 10.3181/00379727-73-17557. [DOI] [PubMed] [Google Scholar]
  10. Olden K., Pratt R. M., Yamada K. M. Role of carbohydrates in protein secretion and turnover: effects of tunicamycin on the major cell surface glycoprotein of chick embryo fibroblasts. Cell. 1978 Mar;13(3):461–473. doi: 10.1016/0092-8674(78)90320-3. [DOI] [PubMed] [Google Scholar]
  11. Robbins P. W., Krag S. S., Liu T. Effects of UDP-glucose addition on the synthesis of mannosyl lipid-linked oligosaccharides by cell-free fibroblast preparations. J Biol Chem. 1977 Mar 10;252(5):1780–1785. [PubMed] [Google Scholar]
  12. Schneider E. G., Nguyen H. T., Lennarz W. J. The effect of tunicamycin, an inhibitor of protein glycosylation, on embryonic development in the sea urchin. J Biol Chem. 1978 Apr 10;253(7):2348–2355. [PubMed] [Google Scholar]
  13. Sefton B. M. Immediate glycosylation of Sindbis virus membrane proteins. Cell. 1977 Apr;10(4):659–668. doi: 10.1016/0092-8674(77)90099-x. [DOI] [PubMed] [Google Scholar]
  14. Speake B. K., Dils R., Mayer R. J. Regulation of enzyme turnover during tissue differention. Studies on the effects of hormones on the turnover of fatty acid synthetase in rabbit mammary gland in organ culture. Biochem J. 1975 May;148(2):309–320. doi: 10.1042/bj1480309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Speake B. K., White D. A. Lipid-linked oligosaccharides containing glucose in lactating rabbit mammary gland. Biochem J. 1978 Dec 15;176(3):993–1000. doi: 10.1042/bj1760993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Speake B. K., White D. A. TThe effect of tunicamycin on glycoprotein biosynthesis in lactating-rabbit mammary gland [proceedings]. Biochem Soc Trans. 1977;5(6):1723–1725. doi: 10.1042/bst0051723. [DOI] [PubMed] [Google Scholar]
  17. Speake B. K., White D. A. The formation of lipid-linked sugars as intermediates in glycoprotein synthesis in rabbit mammary gland. Biochem J. 1978 Feb 15;170(2):273–283. doi: 10.1042/bj1700273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Struck D. K., Lennarz W. J. Evidence for the participation of saccharide-lipids in the synthesis of the oligosaccharide chain of ovalbumin. J Biol Chem. 1977 Feb 10;252(3):1007–1013. [PubMed] [Google Scholar]
  19. TREVELYAN W. E., PROCTER D. P., HARRISON J. S. Detection of sugars on paper chromatograms. Nature. 1950 Sep 9;166(4219):444–445. doi: 10.1038/166444b0. [DOI] [PubMed] [Google Scholar]
  20. Tabas I., Schlesinger S., Kornfeld S. Processing of high mannose oligosaccharides to form complex type oligosaccharides on the newly synthesized polypeptides of the vesicular stomatitis virus G protein and the IgG heavy chain. J Biol Chem. 1978 Feb 10;253(3):716–722. [PubMed] [Google Scholar]
  21. Takatsuki A., Arima K., Tamura G. Tunicamycin, a new antibiotic. I. Isolation and characterization of tunicamycin. J Antibiot (Tokyo) 1971 Apr;24(4):215–223. doi: 10.7164/antibiotics.24.215. [DOI] [PubMed] [Google Scholar]
  22. Takatsuki A., Tamura G. Tunicamycin, a new antibiotic. II. Some biological properties of the antiviral activity of tunicamycin. J Antibiot (Tokyo) 1971 Apr;24(4):224–231. [PubMed] [Google Scholar]
  23. Tkacz J. S., Lampen O. Tunicamycin inhibition of polyisoprenyl N-acetylglucosaminyl pyrophosphate formation in calf-liver microsomes. Biochem Biophys Res Commun. 1975 Jul 8;65(1):248–257. doi: 10.1016/s0006-291x(75)80086-6. [DOI] [PubMed] [Google Scholar]
  24. Waechter C. J., Lennarz W. J. The role of polyprenol-linked sugars in glycoprotein synthesis. Annu Rev Biochem. 1976;45:95–112. doi: 10.1146/annurev.bi.45.070176.000523. [DOI] [PubMed] [Google Scholar]
  25. White D. A. The formation of lipid-linked sugars by cell-free preparations of lactating rabbit mammary gland. Biochem J. 1978 Mar 15;170(3):479–486. doi: 10.1042/bj1700479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. White D. A., Waechter C. J. A mannosyl-carrier lipid of bovine adrenal meddulla and rat parotid. Biochem J. 1975 Mar;146(3):645–651. doi: 10.1042/bj1460645. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES