Processing of O-linked Glycosylation in the Chimera Consisting of α-Subunit and Carboxyl-terminal Peptide of the Human Chorionic Gonadotropin β-Subunit is Affected by Dimer Formation with Follicle-stimulating Hormone β-Subunit

Abstract

hCG, LH, FSH, and TSH are a family of heterodimeric glycoprotein hormones that contain a common α-subunit, but differ in their hormone-specific β-subunits. hCGβ is unique among β-subunits due to a carboxyl-terminal peptide (CTP) bearing four O-linked oligosaccharides. We previously reported that there were differences in O-glycosylation between two chimeras consisting of α-subunit and CTP, i.e. a variant with CTP at the N-terminal region (Cα) and another analog with CTP at the C-terminus (αC) of the α-subunit. To address whether O-glycosylation is influenced by the heterodimer formation, Cα and αC were expressed alone or with FSHβ-subunit in Chinese hamster ovary cells. The O-linked glycosylation was assessed by continuous labeling with [ 35S]methionine/cysteine, immunoprecipitation with anti-α or anti-FSH serum, serial digestion with endoglycosidase-F and neuraminidase, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The decrease in molecular weight of dimeric chimeras digested with endoglycosidase-F was greater in Cα than that in αC after treatment with neuraminidase, revealing that both chimeras have different numbers of sialic acids on O-linked carbohydrates. By treating with endoglycosidase-F, the dimeric αC migrated faster than its free form, whereas the mobility difference between assembled and unassembled forms of Cα was very little. These data indicate that processing of O-glycosylation is affected by the backbone polypeptide chain(s).