Characterization of the Glycan Structures of Glycoprotein GA733-Fc Expressed in a Baculovirus-Insect Cell System

Abstract

Baculovirus-insect cell systems have been used to express functional recombinant biopharmaceutical proteins. Two pFastBac Dual vectors carrying the gene encoding antigen GA733, a cell-surface glycoprotein, fused to the IgG Fc (GA733-Fc) or KDEL (endoplasmic reticulum [ER] retention sequence) (GA733-FcK) genes were constructed to generate baculoviruses expressing the corresponding recombinant proteins in insect cells. The expression of the recombinant GA733-Fc and GA733-FcK proteins and their glycan structure profiles were assessed under various conditions by analyzing the cell line (Sf9 and High Five), the postinfection (PI) time (48, 72, and 96 h), and the harvested sample (cell culture media [CM] or cell lysate [CL]). Immunoblotting showed nonidentical expression of both GA733-Fc and GA733-FcK under various conditions. Glycosylation analysis revealed that varying conditions affected the glycan structure profiles. These results suggest that the PI time, subcellular localization, and cell type affect recombinant protein expression as well as glycosylation in the baculovirus-insect cell system.