Production of recombinant eCG with a potent FSH-like activity

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Equine chorionic gonadotropin (eCG) consists of highly glycosylated alpha-and beta-subunits and belongs to the member of the glycoprotein hormone family which includes luteinizing hormone (LH) and follicle-stimulating hormone (FSH). eCG is a unique member of the gonadotropin family because it elicits response characteristics of both FSH and LH in others species than the horse. The correct conformation of the heterodimer is also important for efficient secretion, hormone-specific post-translational modifications, receptor binding and signal transduction. As an approach to produce recombinant eCG with a potent FSH-like activity, we analyzed the roles of N-linked oligosaccharides in the biological activities of eCG and found that the FSH-like activity of eCG is resistant to the removal of N-linked oligosaccharides. We also investigated whether alpha and beta subunits can be synthesized as a single polypeptide chain (tethered-eCG) with any biological activities by using the tethered-eCG molecule in which the carboxyl terminus of the eCG beta-subunit was fused to the amino terminus of the alpha-subunit. The tethered-eCG was efficiently secreted from the transiently transfected CHO-K1 cells and showed similar LH-like activity to the dimeric eCG. Interestingly, the FSH-like activity of the tethered-eCG was increased markedly in comparison with the native and wild type eCG. Our data for the first time suggest that the tethered-eCG can be expressed efficiently in the CHO-K1 cells without losing LH-and FSH-like activities. Our results also suggest that the recombinant eCG protein with a potent FSH-like activity can possibly be pruduced by E. coli in which very little of the glycosylation occurs.