COMPARISON OF OLIGOSACCHARIDE PROCESSING AMONG VARIOUS INSECT-CELL LINES EXPRESSING A SECRETED GLYCOPROTEIN

The processing of the N-linked oligosaccharide modifying a secreted al kaline phosphatase glycoprotein (SEAP) expressed with a recombinant Au tographa californica nuclear polyhedrosis virus was evaluated in insec t cell lines established from Spodoptera frugiperda, Trichoplusia ni, and Mamestra brassicae. Studies with Endoglycosidase H (Endo H), which removes high-mannose oligosaccharides, revealed that 79% of the intra cellular SEAP produced in the M. brassicae-derived MB0503 cell line wa s Endo H resistant. The commonly used S. frugiperda Sf21 and Sf9 cell lines produced 44 and 21% Endo H-resistant intracellular SEAP, respect ively. Detection of oligosaccharide moieties with lectins, which selec tively recognize terminal sugars, identified only mannose residues on SEAP expressed in the six insect cell lines. However, the oligosacchar ide moiety of SEAP expressed in a Chinese hamster ovary cell line cont ained sialic acid. Therefore, when expressed in mammalian cells, the o ligosaccharide present on SEAP is processed into complex oligosacchari de, but in insect cells it is of the high-mannose type. Studies with i nhibitors of the initial oligosaccharide processing steps demonstrated that all six cell lines possessed glycosidase I/II and mannosidase I activity and that glycosylation was required for secretion.