Study of the sugar chains of recombinant human amyloid precursor protein produced by Chinese hamster ovary cells.
The N- and O-glycans of recombinant amyloid precursor protein (APP), purified from Chinese hamster ovary cells transfected with the human 695-amino acid form of APP, were separately released by hydrazinolysis under different conditions. The reducing ends of the released N- and O-glycans were reduced with NaB3H4 and derivatized with 2-aminobenzamide (2AB), respectively. After acidic N-glycans were obtained by anion-exchange column chromatography, these were converted to neutral oligosaccharides by sialidase digestion, demonstrating that their acidic nature was entirely due to sialylation. The sialidase-treated N-glycans were then fractionated by lectin column chromatography and their structures were determined by linkage-specific sequential exoglycosidase digestion. These results demonstrated that recombinant APP has bi- and triantennary complex type N-glycans with fucosylated and nonfucosylated trimannosyl cores. In a similar fashion, the 2AB-labeled O-glycans derived from APP were determined to be mono- and disialylated core type 1 structures. Taken together, these results indicate that recombinant APP has sialylated bi- and triantennary N-glycans with fucosylated and nonfucosylated cores and sialylated O-glycans with core type 1 structures.
Sato
Y
Y
Department of Glycobiology, Tokyo Metropolitan Institute of Gerontology, Japan.
Liu
C
C
Wojczyk
B S
BS
Kobata
A
A
Spitalnik
S L
SL
Endo
T
T
D016564:Amyloid beta-Protein Precursor / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002853:Chromatography, Liquid / Q000379:methods; D006224:Cricetinae; D003412:Cricetulus; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification
D016564:Amyloid beta-Protein Precursor; D011134:Polysaccharides; D011994:Recombinant Proteins