Biotechnology and Bioprocess Engineering 2019; 24(5): 745-753  
Heterologous Production and Glycosylation of Japanese Eel Follitropin Using Silkworm
Sun Mee Hong1,*, Ji-Hyun Choi1, Sun-Jung Jo1, Kwan-Sik Min2, Dae-Jung Kim3, Jae Man Lee4, and Takahiro Kusakabe4
1Research and Development Department, Gyeongbuk Institute for Marine Bioindustry, Uljin 36315, Korea
2Animal Biotechnology, Graduate School of Future Convergence Technology, Institute of Genetic Engineering, Hankyong National University, Anseong 17579, Korea
3Jeju Fisheries Research Division, National Institute of Fisheries Science, Jeju 63068, Korea
4Laboratory of Insect Genome Science, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, Japan
Correspondence to: Sun Mee Hong
Research and Development Department, Gyeongbuk Institute for Marine Bioindustry, Uljin 36315, Korea
Tel: +82-54-780-3461; Fax: +82-54-780-3469
E-mail: hongsunmee@gimb.or.kr
Received: February 7, 2019; Revised: May 2, 2019; Accepted: July 6, 2019; Published online: October 31, 2019.
© The Korean Society for Biotechnology and Bioengineering. All rights reserved.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
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.
Keywords: Japanese eel, follitropin, silkworm, recombinant protein, glycosylation


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