The glycan processing and site occupancy of recombinant Thy-1 is markedly affected by the presence of a glycosylphosphatidylinositol anchor

• Published in: Glycobiology (Oxford) Vol. 9; no. 12; pp. 1381 - 1387
• Main Authors: Devasahayam, Mercy, Catalino, Peter D., Rudd, Pauline M., Dwek, Raymond A., Barclay, A. Neil
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01.12.1999; Oxford Publishing Limited (England)
• Subjects: Animals; Binding Sites; CHO Cells; Cricetinae; Fucose - analysis; Glycosylation; glycosylphosphatidylinositol anchor; Glycosylphosphatidylinositols - genetics; Glycosylphosphatidylinositols - metabolism; Mannose - analysis; N-Acetylneuraminic Acid - analysis; Rats; Recombinant Proteins - metabolism; sequon occupancy; Solubility; Thy-1; Thy-1 Antigens - chemistry; Thy-1 Antigens - genetics; Thy-1 Antigens - metabolism; Tunicamycin - pharmacology
• ISSN: 0959-6658; 1460-2423
• DOI: 10.1093/glycob/9.12.1381

Thy-1 is a cell surface glycoprotein containing three N-linked glycosylation sites and a glycosylphosphatidylinositol (GPI) anchor. The effect of the anchor on its N-linked glycosylation was investigated by comparing the glycosylation of soluble recombinant Thy-1 (sThy-1) with that of recombinant GPI anchored Thy-1, both expressed in Chinese hamster ovary cells. The sThy-1 was produced in a variety of isoforms including some which lacked carbohydrate on all three sequons whereas the GPI anchored form appeared fully glycosylated like native Thy-1. This was surprising as the attachment of N-linked sugars occurs cotranslationally and it was not expected that the presence of a C-terminal GPI anchor signal sequence would affect sequon occupancy. Furthermore sThy-1 lacking glycosylation could be produced with the inhibitor tunicamycin but in contrast cell surface expression of unglycosylated GPI anchored Thy-1 could not be obtained. The GPI anchored form appeared less processed with almost 4-fold more oligomannose oligosaccharides than in sThy-1 and also with less sialylated and core fucosylated biantennary glycans. Possible mechanisms whereby the anchor or the anchor signal sequence, control site occupancy and maturation are discussed.