Title :
Oligosaccharide processing in the expression of human
plasminogen cDNA by lepidopteran insect (Spodoptera frugiperda) cells
Abstract :
- A comparison has been made between the Asn289-linked oligosaccharide structures Asn289-linked oligosaccharide structures of human plasma plasminogen and a recombinant human plasminogen , expressed in lepidopteran insect (Spodoptera frugiperda) cells, after infection of these cells with a recombinant baculovirus containing the entire human plasminogen cDNA
- Using anion-exchange liquid chromatography mapping of the oligosaccharide units cleaved from the proteins by glycopeptidase F , compared with elution positions of standard oligosaccharide structures , coupled with monosaccharide com position al analysis, we find that the human plasma protein contained only bisialo-biantennary complex-type carbohydrate and asialo-biantennary complex carbohydrate , confirming earlier work published by this laboratory
- The glycosylation pattern of the insect cell expressed recombinant human plasminogen showed considerable microheterogeneity, with identifiable high-mannose carbohydrate (Man9GlcNAc2) and truncated high-mannose oligosaccharide (Man5GlcNAc2, Man4GlcNAc2, and Man3GlcNAc2).
- Of major importance, approximately 40% of the oligosaccharide population consisted of complex carbohydrate (bisialo-biantennary), identical in structure with that of the human plasma protein
- This is the first direct identification of complex carbohydrate in proteins produced in insect cells and demonstrates that trimming and processing of high-mannose carbohydrate into complex-type oligosaccharide can occur
- Our data indicate that both normal and alternate pathways exist in these cells for incorporation and trimming of high-mannose oligosaccharides and that mannosidases, as well as galactosyl-, hexosaminidasyl-, and sialyltransferases are present, and/or can be induced, in these cells
- From these observations, we conclude that amino acid sequences and/or protein conformational properties can control oligosaccharide processing events