Heterologous Production and Glycosylation of Japanese Eel Follitropin Using Silkworm

Abstract

Follitropin, an important gonadotropin hormone, participates in vitellogenesis and spermatogenesis. Equine chorionic gonadotropin (eCG) can induce gonadotropin hormone activity in non-equid species and exhibits a long biological half-life. Here, we report the production, using silkworm larval and pupal systems, of biologically active recombinant hybrid-type follitropins based on the coding sequence of the eCG C-terminal peptide (CTP) between the mature β- and α-chains of eel. The three constructs, rJeFSH, rJeFSH·eCG, and rJeFSH·2xeCG were produced and verified to be N- or O-glycosylated and secreted mature peptides. Although rJeFSH·eCG contains more elaborate O-linked carbohydrate chains than rJeFSH, it elicited no significant in vitro oocyte maturation, which may be a result of insufficient terminal sialylation of its N-and O-linked carbohydrate chains. Then, a hybrid of rJeFSH·2xeCG extended with two eCG CTP. Furthermore, the receptor binding assay revealed potency of rJeFSH and rJeFSH·2xeCG to be a few folds greater than that of rJeFSH·eCG. The findings of this study will be useful for the development of more efficient GTHs in teleosts, including eels, when various modifications with two or more extended eCG CTP produced by silkworm are included.