Glycoproteins secreted from suspension-cultured tobacco BY2 cells have distinct glycan structures from intracellular glycoproteins

Glycan structures of glycoproteins secreted in the spent medium of tobacco BY2 suspension-cultured cells,were analyzed. The N-glycans were liberated b y hydrazinolysis and the resulting oligosaccharides were labeled with 2-ami nopyridine. The pyridylaminated (PA) glycans were purified by reversed-phas e and size-fractionation HPLC. The structures of the PA sugar chains were i dentified by a combination of the two-dimensional PA sugar chain mapping, M S analysis, and exoglycosidase digestion. The ratio (40:60) of the amount o f glycans with high-mannose-type structure to that with plant-complex-type structure of extracellular glycoproteins is significantly different from th at (ratio 10:90) previously found in intracellular glycoproteins [Palacpac et al., Biosci. Biotechnol. Biochem. 63 (1999) 35-39]. Extracellular glycop roteins have six distinct N-glycans (marked by *) from intracellular glycop roteins, and the high-mannose-type structures account for nearly, 40% (Man( 5)GlcNac(2), 28.8%; Man(6)GlcNAc(2)*, 6.4%; and Man(7)GIcNAc(2)*, 3.8%), wh ile the plant-complex-type structures account for nearly 60% (GlcNAc(2)Man( 3) Xyl(1)GlcNAc(2)*, 32.1%; GlcNAc(1)Man(3)Xyl(1)GlcNAc(2) (containing two isomers)*, 6.2%, GlcNAc(2)Man(3) GlcNAc(2)*, 4.9%; Man(3)Xyl(1)Fuc(1)GlcNAc (2), 8.3%; and Man(3)Xyl(1)GlcNAc(2), 3.7%).