Biomaterial Database

N-Linked glycosylation of a baculovirus-expressed recombinant glycoprotein in insect larvae and tissue culture cells. 1998

P C Kulakosky, and P R Hughes, and H A Wood

Boyce Thompson Institute for Plant Research, Tower Road, Ithaca, NY 14853-1801, USA.

The potential of insect cell cultures and larvae infected with recombinant baculoviruses to produce authentic recombinant glycoproteins cloned from mammalian sources was investigated. A comparison was made of the N-linked glycans attached to secreted alkaline phosphatase (SEAP) produced in four species of insect larvae and their derived cell lines plus one additional insect cell line and larvae of one additional species. These data survey N-linked oligosaccharides produced in four families and six genera of the order Lepidoptera. Recombinant SEAP expressed by recombinant isolates of Autographa californica and Bombyx mori nucleopolyhedroviruses was purified from cell culture medium, larval hemolymph or larval homogenates by phosphate affinity chromatography. The N-linked oligosaccharides were released with PNGase-F, labeled with 8-aminonaphthalene-1-3-6-trisulfonic acid, fractionated by polyacrylamide gel electrophoresis, and analyzed by fluorescence imaging. The oligosaccharide structures were confirmed with exoglycosidase digestions. Recombinant SEAP produced in cell lines of Lymantria dispar (IPLB-LdEIta), Heliothis virescens (IPLB-HvT1), and Bombyx mori (BmN) and larvae of Spodoptera frugiperda, Trichoplusia ni , H.virescens , B.mori , and Danaus plexippus contained oligosaccharides that were structurally identical to the 10 oligosaccharides attached to SEAP produced in T.ni cell lines. The oligosaccharide structures were all mannose-terminated. Structures containing two or three mannose residues, with and without core fucosylation, constituted more than 75% of the oligosaccharides from the cell culture and larval samples.

UI	MeSH Term	Description	Entries
D007814	Larva	Wormlike or grublike stage, following the egg in the life cycle of insects, worms, and other metamorphosing animals.	Maggots,Tadpoles,Larvae,Maggot,Tadpole
D007915	Lepidoptera	A large order of insects comprising the butterflies and moths.
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D009844	Oligosaccharides	Carbohydrates consisting of between two (DISACCHARIDES) and ten MONOSACCHARIDES connected by either an alpha- or beta-glycosidic link. They are found throughout nature in both the free and bound form.	Oligosaccharide
D011994	Recombinant Proteins	Proteins prepared by recombinant DNA technology.	Biosynthetic Protein,Biosynthetic Proteins,DNA Recombinant Proteins,Recombinant Protein,Proteins, Biosynthetic,Proteins, Recombinant DNA,DNA Proteins, Recombinant,Protein, Biosynthetic,Protein, Recombinant,Proteins, DNA Recombinant,Proteins, Recombinant,Recombinant DNA Proteins,Recombinant Proteins, DNA
D002240	Carbohydrate Sequence	The sequence of carbohydrates within POLYSACCHARIDES; GLYCOPROTEINS; and GLYCOLIPIDS.	Carbohydrate Sequences,Sequence, Carbohydrate,Sequences, Carbohydrate
D002460	Cell Line	Established cell cultures that have the potential to propagate indefinitely.	Cell Lines,Line, Cell,Lines, Cell
D006023	Glycoproteins	Conjugated protein-carbohydrate compounds including MUCINS; mucoid, and AMYLOID glycoproteins.	C-Glycosylated Proteins,Glycosylated Protein,Glycosylated Proteins,N-Glycosylated Proteins,O-Glycosylated Proteins,Glycoprotein,Neoglycoproteins,Protein, Glycosylated,Proteins, C-Glycosylated,Proteins, Glycosylated,Proteins, N-Glycosylated,Proteins, O-Glycosylated
D006031	Glycosylation	The synthetic chemistry reaction or enzymatic reaction of adding carbohydrate or glycosyl groups. GLYCOSYLTRANSFERASES carry out the enzymatic glycosylation reactions. The spontaneous, non-enzymatic attachment of reducing sugars to free amino groups in proteins, lipids, or nucleic acids is called GLYCATION (see MAILLARD REACTION).	Protein Glycosylation,Glycosylation, Protein
D000469	Alkaline Phosphatase	An enzyme that catalyzes the conversion of an orthophosphoric monoester and water to an alcohol and orthophosphate. EC 3.1.3.1.