Title : Carbohydrate structure of recombinant human uterine
tissue plasminogen activator expressed in mouse epithelial cells
Abstract :
- Recombinant human uterine tissue plasminogen activator ( tPA ), in part metabolically labeled with [6-3H]glucosamine or [35S]sulfate, was isolated from mouse epithelial cells (C127)
- Oligosaccharides present were liberated by treatment of tryptic glycopeptides with endo- beta-N-acetylglucosaminidase H or peptide-N4-(N-acetyl-beta-glucosaminyl) asparagine amidase F and fractionated by high-performance liquid chromatography
- The glycans were characterized by digestion with exoglycosidases, methylation analysis and, in part, by acetolysis and 1H-NMR spectroscopy
- Glycopeptides comprising individual glycosylation sites were identified by N-terminal amino acid sequencing
- The results demonstrate that recombinant tPA from C127 cells carries at Asn117 oligomannosidic glycans with 5-8 mannose residues as well as small amounts of hybrid-type species
- Asn184 is only partially glycosylated and substituted by fucosylated triantennary and small amounts of diantennary N-acetyllactosaminic glycans
- Likewise, Asn448 carries predominantly fucosylated triantennary species, in addition to, small amounts of diantennary and tetraantennary oligosaccharides
- As a characteristic feature, part of the triantennary glycans at Asn184 and Asn448 contain additional Gal(alpha 1-3) substituents and/or sulfate groups linked to position six of beta-galactosyl residues forming NeuAc(alpha 2-3)[ HO3S-6]Gal ( beta 1-4 ) units
- Oligosaccharides attached to Asn448 are almost completely substituted by (alpha 2-3)- or (alpha 2-6)-linked sialic acid residues and carry the majority of sulfate groups present
- Glycans at Asn184 were found to be less sialylated and sulfated
Output (sent_index, trigger,
protein,
sugar,
site):
- 0. activator, , tissue plasminogen activator, Carbohydrate structure, -
- 10. Asn184, , -, Glycans, Asn184
- 10. sialylated, , -, Glycans, -
- 2. glycopeptides, , -, -, glycopeptides
- 4. glycosylation, , -, -, sites
- 5. Asn117, , -, Asn117 oligomannosidic glycans, Asn117
- 6. fucosylated, , -, diantennary N-acetyllactosaminic glycans, -
- 6. glycosylated, , -, -, Asn184
- 8. Asn184, , -, the triantennary glycans, Asn184 and Asn448
- 8. Asn448, , -, the triantennary glycans, Asn184 and Asn448
- 8. linked, , -, additional Gal(alpha 1-3) substituents, position
- 9. attached, , -, Oligosaccharides, Asn448
Output(Part-Of) (sent_index,
protein,
site):
*Output_Site_Fusion* (sent_index,
protein,
sugar,
site):
- 10. tissue plasminogen activator, Glycans, Asn184
- 5. tissue plasminogen activator, Asn117 oligomannosidic glycans, Asn117
- 6. tissue plasminogen activator, -, Asn184
- 8. tissue plasminogen activator, the triantennary glycans, Asn184 and Asn448
- 9. tissue plasminogen activator, Oligosaccharides, Asn448