Production of Recombinant Single-Chain Eel Luteinizing Hormone and Follicle-Stimulating Hormone Analogs in Chinese Hamster Ovary Suspension Cell Culture
We produced rec-single chain eel luteinizing (rec-eel LH) and follicle-stimulating (rec- eel FSH) hormones displaying high biological activity in Chinese hamster ovary suspension (CHO-S) cells. We constructed several mutants, in which a linker, including an O-linked glycosylated carboxyl-terminal pe...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2024-01-01
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| Series: | Current Issues in Molecular Biology |
| Subjects: | |
| Online Access: | https://www.mdpi.com/1467-3045/46/1/35 |
| Summary: | We produced rec-single chain eel luteinizing (rec-eel LH) and follicle-stimulating (rec- eel FSH) hormones displaying high biological activity in Chinese hamster ovary suspension (CHO-S) cells. We constructed several mutants, in which a linker, including an O-linked glycosylated carboxyl-terminal peptide (CTP) of an equine chorionic gonadotropin (eCG) β-subunit, was attached between the β- and α-subunit (LH-M and FSH-M) or in the N-terminal (C-LH and C-FSH) or C-terminal (LH-C and FSH-C) regions. The plasmids were transfected into CHO-S cells, and culture supernatants were collected. The secretion of mutants from the CHO-S cells was faster than that of eel LHβ/α-wt and FSHβ/α-wt proteins. The molecular weight of eel LHβ/α-wt and eel FSHβ/α-wt was 32–34 and 34–36 kDa, respectively, and that of LH-M and FSH-M was 40–43 and 42–45 kDa, respectively. Peptide-N-glycanase F-treatment markedly decreased the molecular weight by approximately 8–10 kDa. The EC<sub>50</sub> value and the maximal responsiveness of the eel LH-M and eel FSH-M increased compared with the wild-type proteins. These results show that the CTP region plays a pivotal role in early secretion and signal transduction. We suggest that novel rec-eel LH and FSH proteins, exhibiting potent activity, could be produced in large quantities using a stable CHO cell system. |
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| ISSN: | 1467-3037 1467-3045 |