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. 1990 Dec;9(13):4249–4258. doi: 10.1002/j.1460-2075.1990.tb07873.x

Biosynthesis of glycosyl-phosphatidylinositol lipids in Trypanosoma brucei: involvement of mannosyl-phosphoryldolichol as the mannose donor.

A K Menon 1, S Mayor 1, R T Schwarz 1
PMCID: PMC552207  PMID: 2148289

Abstract

Trypanosome variant surface glycoproteins (VSGs) exemplify a class of eukaryotic cell-surface glycoproteins that rely on a covalently attached lipid, glycosyl-phosphatidylinositol, for membrane attachment. The glycolipid anchor is acquired soon after translation of the polypeptide, apparently by replacement of a short sequence of carboxyl-terminal amino acids with a precursor glycolipid. A candidate glycolipid precursor (P2) and a related glycolipid (P3) have been identified in polar lipid extracts from trypanosomes. Both lipids are glycosylphosphatidylinositol species containing a Man3GlcN core glycan indistinguishable from the backbone sequence of the VSG glycolipid anchor. We and others have recently described the cell-free synthesis of P2, P3, and a spectrum of putative biosynthetic lipid intermediates using crude preparations of trypanosome membranes. In this paper we use these preparations to show that all three mannose residues in the glycosyl-phosphatidylinositol glycan are derived from dolichol-P-mannose.

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Selected References

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  1. Banerjee D. K. Amphomycin inhibits mannosylphosphoryldolichol synthesis by forming a complex with dolichylmonophosphate. J Biol Chem. 1989 Feb 5;264(4):2024–2028. [PubMed] [Google Scholar]
  2. Banerjee D. K., Scher M. G., Waechter C. J. Amphomycin: effect of the lipopeptide antibiotic on the glycosylation and extraction of dolichyl monophosphate in calf brain membranes. Biochemistry. 1981 Mar 17;20(6):1561–1568. doi: 10.1021/bi00509a024. [DOI] [PubMed] [Google Scholar]
  3. Belard M., Cacan R., Verbert A. Characterization of an oligosaccharide-pyrophosphodolichol pyrophosphatase activity in yeast. Biochem J. 1988 Oct 1;255(1):235–242. [PMC free article] [PubMed] [Google Scholar]
  4. Chapman A., Fujimoto K., Kornfeld S. The primary glycosylation defect in class E Thy-1-negative mutant mouse lymphoma cells is an inability to synthesize dolichol-P-mannose. J Biol Chem. 1980 May 25;255(10):4441–4446. [PubMed] [Google Scholar]
  5. Chapman A., Trowbridge I. S., Hyman R., Kornfeld S. Structure of the lipid-linked oligosaccharides that accumulate in class E Thy-1-negative mutant lymphomas. Cell. 1979 Jul;17(3):509–515. doi: 10.1016/0092-8674(79)90259-9. [DOI] [PubMed] [Google Scholar]
  6. Chojnacki T., Dallner G. The biological role of dolichol. Biochem J. 1988 Apr 1;251(1):1–9. doi: 10.1042/bj2510001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Conzelmann A., Spiazzi A., Hyman R., Bron C. Anchoring of membrane proteins via phosphatidylinositol is deficient in two classes of Thy-1 negative mutant lymphoma cells. EMBO J. 1986 Dec 1;5(12):3291–3296. doi: 10.1002/j.1460-2075.1986.tb04642.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Datema R., Schwarz R. T., Winkler J. Glycosylation of influenza virus proteins in the presence of fluoroglucose occurs via a different pathway. Eur J Biochem. 1980 Sep;110(2):355–361. doi: 10.1111/j.1432-1033.1980.tb04875.x. [DOI] [PubMed] [Google Scholar]
  9. Doering T. L., Masterson W. J., Englund P. T., Hart G. W. Biosynthesis of the glycosyl phosphatidylinositol membrane anchor of the trypanosome variant surface glycoprotein. Origin of the non-acetylated glucosamine. J Biol Chem. 1989 Jul 5;264(19):11168–11173. [PubMed] [Google Scholar]
  10. Doering T. L., Masterson W. J., Hart G. W., Englund P. T. Biosynthesis of glycosyl phosphatidylinositol membrane anchors. J Biol Chem. 1990 Jan 15;265(2):611–614. [PubMed] [Google Scholar]
  11. Elbein A. D. Inhibitors of the biosynthesis and processing of N-linked oligosaccharide chains. Annu Rev Biochem. 1987;56:497–534. doi: 10.1146/annurev.bi.56.070187.002433. [DOI] [PubMed] [Google Scholar]
  12. Fatemi S. H., Haas R., Jentoft N., Rosenberry T. L., Tartakoff A. M. The glycophospholipid anchor of Thy-1. Biosynthetic labeling experiments with wild-type and class E Thy-1 negative lymphomas. J Biol Chem. 1987 Apr 5;262(10):4728–4732. [PubMed] [Google Scholar]
  13. Fatemi S. H., Tartakoff A. M. Hydrophilic anchor-deficient Thy-1 is secreted by a class E mutant T lymphoma. Cell. 1986 Aug 29;46(5):653–657. doi: 10.1016/0092-8674(86)90340-5. [DOI] [PubMed] [Google Scholar]
  14. Ferguson M. A., Williams A. F. Cell-surface anchoring of proteins via glycosyl-phosphatidylinositol structures. Annu Rev Biochem. 1988;57:285–320. doi: 10.1146/annurev.bi.57.070188.001441. [DOI] [PubMed] [Google Scholar]
  15. Frahm R., Schwarz M., Geishauser M. Seltene Hypersupinationsluxation im distalen radioulnaren Gelenk. Rofo. 1989 Jun;150(6):746–748. doi: 10.1055/s-2008-1047119. [DOI] [PubMed] [Google Scholar]
  16. Haselbeck A., Tanner W. Dolichyl phosphate-mediated mannosyl transfer through liposomal membranes. Proc Natl Acad Sci U S A. 1982 Mar;79(5):1520–1524. doi: 10.1073/pnas.79.5.1520. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Haselbeck A., Tanner W. O-glycosylation in Saccharomyces cerevisiae is initiated at the endoplasmic reticulum. FEBS Lett. 1983 Jul 25;158(2):335–338. doi: 10.1016/0014-5793(83)80608-5. [DOI] [PubMed] [Google Scholar]
  18. Higgins J. A., Hitchin B. W., Low M. G. Phosphatidylinositol-specific phospholipase C of Bacillus thuringiensis as a probe for the distribution of phosphatidylinositol in hepatocyte membranes. Biochem J. 1989 May 1;259(3):913–916. doi: 10.1042/bj2590913. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hirschberg C. B., Snider M. D. Topography of glycosylation in the rough endoplasmic reticulum and Golgi apparatus. Annu Rev Biochem. 1987;56:63–87. doi: 10.1146/annurev.bi.56.070187.000431. [DOI] [PubMed] [Google Scholar]
  20. Hull S. R., Turco S. J. Separation of partially methylated mannitols by liquid chromatography. Anal Biochem. 1985 Dec;151(2):554–560. doi: 10.1016/0003-2697(85)90219-2. [DOI] [PubMed] [Google Scholar]
  21. Hyman R. Cell-surface-antigen mutants of haematopoietic cells. Tools to study differentiation, biosynthesis and function. Biochem J. 1985 Jan 1;225(1):27–40. doi: 10.1042/bj2250027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kang M. S., Spencer J. P., Elbein A. D. Amphomycin inhibition of mannose and GlcNAc incorporation into lipid-linked saccharides. J Biol Chem. 1978 Dec 25;253(24):8860–8866. [PubMed] [Google Scholar]
  23. Krag S. S., Robbins P. W. Sindbis envelope proteins as endogenous acceptors in reactions of guanosine diphosphate-[14C]Mannose with preparations of infected chicken embryo fibroblasts. J Biol Chem. 1977 Apr 25;252(8):2621–2629. [PubMed] [Google Scholar]
  24. Krakow J. L., Doering T. L., Masterson W. J., Hart G. W., Englund P. T. A glycolipid from Trypanosoma brucei related to the variant surface glycoprotein membrane anchor. Mol Biochem Parasitol. 1989 Oct;36(3):263–270. doi: 10.1016/0166-6851(89)90174-6. [DOI] [PubMed] [Google Scholar]
  25. Krakow J. L., Hereld D., Bangs J. D., Hart G. W., Englund P. T. Identification of a glycolipid precursor of the Trypanosoma brucei variant surface glycoprotein. J Biol Chem. 1986 Sep 15;261(26):12147–12153. [PubMed] [Google Scholar]
  26. Lennarz W. J. Protein glycosylation in the endoplasmic reticulum: current topological issues. Biochemistry. 1987 Nov 17;26(23):7205–7210. doi: 10.1021/bi00397a001. [DOI] [PubMed] [Google Scholar]
  27. Masterson W. J., Doering T. L., Hart G. W., Englund P. T. A novel pathway for glycan assembly: biosynthesis of the glycosyl-phosphatidylinositol anchor of the trypanosome variant surface glycoprotein. Cell. 1989 Mar 10;56(5):793–800. doi: 10.1016/0092-8674(89)90684-3. [DOI] [PubMed] [Google Scholar]
  28. Masterson W. J., Raper J., Doering T. L., Hart G. W., Englund P. T. Fatty acid remodeling: a novel reaction sequence in the biosynthesis of trypanosome glycosyl phosphatidylinositol membrane anchors. Cell. 1990 Jul 13;62(1):73–80. doi: 10.1016/0092-8674(90)90241-6. [DOI] [PubMed] [Google Scholar]
  29. Mayor S., Menon A. K., Cross G. A., Ferguson M. A., Dwek R. A., Rademacher T. W. Glycolipid precursors for the membrane anchor of Trypanosoma brucei variant surface glycoproteins. I. Can structure of the phosphatidylinositol-specific phospholipase C sensitive and resistant glycolipids. J Biol Chem. 1990 Apr 15;265(11):6164–6173. [PubMed] [Google Scholar]
  30. Mayor S., Menon A. K., Cross G. A. Glycolipid precursors for the membrane anchor of Trypanosoma brucei variant surface glycoproteins. II. Lipid structures of phosphatidylinositol-specific phospholipase C sensitive and resistant glycolipids. J Biol Chem. 1990 Apr 15;265(11):6174–6181. [PubMed] [Google Scholar]
  31. Menon A. K., Mayor S., Ferguson M. A., Duszenko M., Cross G. A. Candidate glycophospholipid precursor for the glycosylphosphatidylinositol membrane anchor of Trypanosoma brucei variant surface glycoproteins. J Biol Chem. 1988 Feb 5;263(4):1970–1977. [PubMed] [Google Scholar]
  32. Menon A. K., Schwarz R. T., Mayor S., Cross G. A. Cell-free synthesis of glycosyl-phosphatidylinositol precursors for the glycolipid membrane anchor of Trypanosoma brucei variant surface glycoproteins. Structural characterization of putative biosynthetic intermediates. J Biol Chem. 1990 Jun 5;265(16):9033–9042. [PubMed] [Google Scholar]
  33. Palamarczyk G., Lehle L., Mankowski T., Chojnacki T., Tanner W. Specificity of solubilized yeast glycosyl transferases for polyprenyl derivatives. Eur J Biochem. 1980 Apr;105(3):517–523. doi: 10.1111/j.1432-1033.1980.tb04527.x. [DOI] [PubMed] [Google Scholar]
  34. Perez M., Hirschberg C. B. Translocation of UDP-N-acetylglucosamine into vesicles derived from rat liver rough endoplasmic reticulum and Golgi apparatus. J Biol Chem. 1985 Apr 25;260(8):4671–4678. [PubMed] [Google Scholar]
  35. Roberts W. L., Myher J. J., Kuksis A., Low M. G., Rosenberry T. L. Lipid analysis of the glycoinositol phospholipid membrane anchor of human erythrocyte acetylcholinesterase. Palmitoylation of inositol results in resistance to phosphatidylinositol-specific phospholipase C. J Biol Chem. 1988 Dec 15;263(35):18766–18775. [PubMed] [Google Scholar]
  36. Roberts W. L., Santikarn S., Reinhold V. N., Rosenberry T. L. Structural characterization of the glycoinositol phospholipid membrane anchor of human erythrocyte acetylcholinesterase by fast atom bombardment mass spectrometry. J Biol Chem. 1988 Dec 15;263(35):18776–18784. [PubMed] [Google Scholar]
  37. Schwarz R. T., Datema R. Inhibition of the dolichol pathway of protein glycosylation. Methods Enzymol. 1982;83:432–443. doi: 10.1016/0076-6879(82)83041-3. [DOI] [PubMed] [Google Scholar]
  38. Sharma C. B., Babczinski P., Lehle L., Tanner W. The role of dolicholmonophosphate in glycoprotein biosynthesis in Saccharomyces cerevisiae. Eur J Biochem. 1974 Jul 1;46(1):35–41. doi: 10.1111/j.1432-1033.1974.tb03594.x. [DOI] [PubMed] [Google Scholar]
  39. Trowbridge I. S., Hyman R. Abnormal lipid-linked oligosaccharides in class E Thy-1-negative mutant lymphomas. Cell. 1979 Jul;17(3):503–508. doi: 10.1016/0092-8674(79)90258-7. [DOI] [PubMed] [Google Scholar]
  40. Vijay I. K., Perdew G. H. Biosynthesis of mammary glycoproteins. Structural characterization of different isomers of lipid-linked hexa- and heptasaccharides. J Biol Chem. 1980 Dec 10;255(23):11221–11226. [PubMed] [Google Scholar]

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