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Sialylation of N-Glycans on the Recombinant Proteins Expressed by a Baculovirus-Insect Cell System under β-N-Acetylglucosaminidase Inhibition

2002; Elsevier BV; Volume: 277; Issue: 7 Linguagem: Inglês

10.1074/jbc.m110548200

1083-351X

Satoko Watanabe, Takehiro Kokuho, Hitomi Takahashi, Masashi Takahashi, Takayuki Kubota, Shigeki Inumaru,

Invertebrate Immune Response Mechanisms

We investigated the ability of a baculovirus-insect cell system to produce sialylated glycoproteins. Despite the presence of enzymes for synthesizing complex-type N -glycans, the most frequent structure of insect N -glycan is the paucimannosidic type, Man 3 GlcNAc 2 (±Fuc). The reason for the overwhelming assembly of paucimannosidic N -glycans is not yet well understood. We hypothesized that this predominance might be due to insect-specific, Golgi-associated β- N -acetylglucosaminidase (GlcNAcase)-mediated removal of N -acetylglucosamine residues from the precursor N -glycan, thereby preventing its galactosylation and terminal sialylation. As we expected, the suppression of intrinsic GlcNAcase activity with a specific inhibitor, 2-acetamido-1,2-dideoxynojirimycin, allowed the accumulation of sialylated glycoproteins in the supernatants of insect cell cultures after baculoviral infection. Our observation indicates that GlcNAcase-dependent depletion of N -acetylglucosamine residues from intermediate N -glycans is critical for the assembly of paucimannosidic N -glycans in insect cells and, more importantly, that insect cells (under specific conditions) retain the ability to construct sialylated N -glycans like those in mammalian cells.