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

Abstract

Recombinant baculoviruses vAcßhCG COR and vAcßhCG POL, carrying the gene ( ßhCG) 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 ßhCG (re-ßhCG) revealed that the secretion of βhCG COR was relatively higher. Enzymatic and chemical analysis of carbohydrates showed that ßhCG COR was more glycosylated than ßhCG POL. However, the insect-derived ßhCG, with a high-mannose type of sugar, was glycosylated differently and to a lesser extent when compared with the native, urinary ßhCG, and consequently, ßhCG COR was more bioactive on a unit-mass basis than ßhCG 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.