N-glycan patterns of human transferrin produced in Trichoplusia ni insect cells: effects of mammalian galactosyltransferase.

Abstract:

:The N-glycans of human serum transferrin produced in Trichopulsia ni cells were analyzed to examine N-linked oligosaccharide processing in insect cells. Metabolic radiolabeling of the intra- and extracellular protein fractions revealed the presence of multiple transferrin glycoforms with molecular weights lower than that observed for native human transferrin. Consequently, the N-glycan structures of transferrin in the culture medium were determined using three-dimensional high performance liquid chromatography. The attached oligosaccharides included high mannose, paucimannosidic, and hybrid structures with over 50% of these structures containing one fucose, alpha(1,6)-, or two fucoses, alpha(1,6)- and alpha(1,3)-, linked to the Asn-linked N-acetylglucosamine. Neither sialic acid nor galactose was detected on any of the N-glycans. However, when transferrin was coexpressed with beta(1,4)-galactosyltransferase three additional galactose-containing hybrid oligosaccharides were obtained. The galactose attachments were exclusive to the alpha(1, 3)-mannose branch and the structures varied by the presence of zero, one, or two attached fucose residues. Furthermore, the presence of the galactosyltransferase appeared to reduce the number of paucimannosidic structures, which suggests that galactose attachment inhibits the ability of hexosaminidase activity to remove the terminal N-acetylglucosamine. The ability to promote galactosylation and reduce paucimannosidic N-glycans suggests that the oligosaccharide processing pathway in insect cells may be manipulated to mimic more closely that of mammalian cells.

journal_name

Glycobiology

journal_title

Glycobiology

authors

Ailor E,Takahashi N,Tsukamoto Y,Masuda K,Rahman BA,Jarvis DL,Lee YC,Betenbaugh MJ

doi

10.1093/glycob/10.8.837

subject

Has Abstract

pub_date

2000-08-01 00:00:00

pages

837-47

issue

8

eissn

0959-6658

issn

1460-2423

journal_volume

10

pub_type

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