N-ACETYL-BETA-GLUCOSAMINIDASE ACCOUNTS FOR DIFFERENCES IN GLYCOSYLATION OF INFLUENZA-VIRUS HEMAGGLUTININ EXPRESSED IN INSECT CELLS FROM A BACULOVIRUS VECTOR

The hemagglutinin of fowl plague virus has been expressed in Spodopter a frugiperda (Sf9) cells and in Estigmene acrea cells by using a bacul ovirus vector. Structural analysis revealed that the endo-H-resistant N-glycans of HA from Sf9 cells were predominantly trimannosyl core oli gosaccharides, whereas in E. ncr ea cells most of these cores were elo ngated by at least one terminal N-acetylglucosamine residue. To unders tand the difference in carbohydrate structures, enzymes involved in N- glycan processing have been analyzed. The results revealed that the di fferent glycosylation patterns observed are due to an N-acetyl-beta-gl ucosaminidase activity that was found in Sf9 cells but not in E. acrea cells. This enzyme specifically used the GlcNAcMan(3)GlcNAc(2) oligos accharide as a substrate. When N-acetyl-beta-glucosaminidase or alpha- mannosidase II was inhibited by specific inhibitors, the amount of ter minal N-acetylglucosamine in hemagglutinin from Sf9 cells was signific antly enhanced. These results demonstrate that N glycosylation in both cell lines follows the classical pathway up to the stage of GlcNAcMan (3)GlcNAc(2) oligosaccharide side chains. Whereas these structures are the end product in E. acrea cells, they are degraded in Sf9 cells to Man(3)GlcNAc(2) cores by N-acetyl-beta-glucosaminidase.