Biological activities of tethered equine chorionic gonadotropin (eCG) and its deglycosylated mutants.
Equine chorionic gonadotropin (eCG), which consists of highly glycosylated alpha- and beta-subunits, is a unique member of the gonadotropin family because it elicits response characteristics of both follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in species other than the horse. In this study, recombinant tethered-eCG as well as its deglycosylated mutants were produced to determine if alpha- and beta- subunits can be synthesized as a single polypeptide chain (tethered-eCG) and display biological activity. We found that tethered-eCG (T- betaalpha) had both LH- and FSH-like activities comparable to dimeric eCG. Luteinizing hormone-like activity of tethered-eCGs deglycosylated at Asn(56) (T-betaalpha56) was decreased. In contrast, LH-like activity of eCG without O-glycosylated carboxyl-terminal peptide (CTP) (T-betacalpha) was slightly decreased but still similar to T-betaalpha. Double mutation at Asn(56) and CTP (T-betacalpha56) caused marked decrease in the activity, indicating that both glycosylations at Asn(56) and CTP are involved in LH-like activity in the tethered form. Interestingly, FSH-like activity remained in all deglycosylated eCG mutants (T-betaalpha56, T-betacalpha and T-betacalpha56) as well as T-betaalpha. The biological roles of oligosaccharides at Asn(56) of eCG alpha-subunit and O-linked peptide of beta-subunit appear to be different in LH- and FSH-like activities in tethered-eCG.
Min
Kwan-Sik
KS
Laboratory of Cellular Biochemistry, Animal Resource Sciences /Veterinary Medical Sciences, Graduate School of Agriculture and Life Sciences, The University of Tokyo, Japan.
Hiyama
Takashi
T
Seong
Hwan-Hoo
HH
Hattori
Naka
N
Tanaka
Satoshi
S
Shiota
Kunio
K
D000818:Animals; D001216:Asparagine / Q000737:chemistry; D001681:Biological Assay; D016466:CHO Cells; D006063:Chorionic Gonadotropin / Q000378:metabolism*; D006224:Cricetinae; D004247:DNA / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005640:Follicle Stimulating Hormone / Q000378:metabolism; D005822:Genetic Vectors; D006031:Glycosylation; D006728:Hormones / Q000737:chemistry; D006736:Horses; D007986:Luteinizing Hormone / Q000737:chemistry; D008957:Models, Genetic; D009154:Mutation; D009844:Oligosaccharides / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D014162:Transfection
D006063:Chorionic Gonadotropin; D006728:Hormones; D009844:Oligosaccharides; D010455:Peptides; D011994:Recombinant Proteins; D001216:Asparagine; D007986:Luteinizing Hormone; D005640:Follicle Stimulating Hormone; D004247:DNA