Human alpha 1-acid glycoprotein as a model protein for glycoanalysis in baculovirus expression vector system

• Recombinant human α1AGP was expressed and highly glycosylated in silkworm cells and larvae using BEVS.
• The N-linked glycan structure of the recombinant human α1AGP was identified as the paucimannosidic type.
• α1AGP is a good model glycoprotein for analyzing the N-glycosylation alterations in insect-BEVS.

Glycosylation is an important post-translational modification that confers various biological activities, structural stability, and inter-molecular interactions to proteins. Baculovirus expression vector system (BEVS) is widely used to produce recombinant glycoproteins, which may not be suitable for clinical use due to differences in the N-linked glycan structure between insects and mammals. It is necessary to develop an appropriate model protein-base platform for glycoanalysis to engineer the insect-type N-glycosylation pathway into human type efficiently. In this study, we employed human plasma protein alpha 1-acid glycoprotein (α1AGP). It was highly secreted from cultured silkworm cells and larvae when using the BEVS and glycosylated with insect type N-linked glycans. Interestingly, when separated on SDS-PAGE, the purified recombinant α1AGP secreted into silkworm haemolymph generated six distinct products from three alternative translates, suggesting that α1AGP has variations for the recognition or choice of glycosylation sites.

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