Differential secretion and glycosylation of recombinant human chorionic gonadotropin (ßhCG) synthesized using different promoters in the baculovirus expression vector system

Abstract

Recombinant baculoviruses vAcβ hCG_COR and vAcβ hCG_POL, carrying the gene (β hCG) encoding the β-subunit of human chorionic gonadotropin under the transcriptional control of the late AcNPV core protein gene promoter (p_cor) and the very late polyhedrin gene promoter (P_POL), respectively, were constructed and used to infect lepidopteran cells. Western blot analysis of intra- and extracellular recombinant β hCG (re-β hCG) revealed that the secretion of βhCG_COR was relatively higher. Enzymatic and chemical analysis of carbohydrates showed that ßhCG_COR was more glycosylated than β hCG_POL. However, the insect-derived β hCG, with a high-mannose type of sugar, was glycosylated differently and to a lesser extent when compared with the native, urinary β hCG, and consequently, β hCG_COR was more bioactive on a unit-mass basis than β hCG_POL. This temporal gene expression strategy, besides being able to circumvent the 'secretory load' encountered during the synthesis of extensively glycosylated proteins in the baculovirus system, also offers a model to study the role of carbohydrates, both in qualitative and quantitative terms, in protein structure and function.