The role of the human chorionic gonadotropin (hCG) N-linked oligosaccharides in receptor binding and signal transduction was analyzed using site-directed mutagenesis and transfection studies. hCG derivatives with alterations at individual glycosylation sites were expressed in Chinese hamster ovary cells. Receptor binding studies showed that absence of any or all of the hCG N-linked oligosaccharides had only a minor effect on the receptor affinity of the derivatives. Similarly, absence of the N-linked oligosaccharides from the β subunit or a single oligosaccharide from Asn-78 of α had no effect on the production of cAMP or on steroidogenesis. However, the absence of carbohydrate at Asn-52 of α decreases both the steroidogenic and cAMP responses. Furthermore, absence of this critical oligosaccharide unit on α unmasks differences in the two N-linked oligosaccharides on β; the β Asn-13 oligosaccharide but not the β Asn-30 oligosaccharide plays a more important role in steroidogenesis. Dimers containing deglycosylated β subunit and an α subunit lacking either the Asn-52 oligosaccharide or both oligosaccharides fail to stimulate cAMP or steroid formation. Moreover, these derivatives bind to receptor and behave as competitive antagonists. The use of site-directed mutagenesis was critical in uncovering site-specific functions of the hCG N-linked oligosaccharides in signal transduction and reveals the importance of the Asn-52 oligosaccharide in this process.
|Number of pages||6|
|Journal||Journal of Biological Chemistry|
|State||Published - Jan 1 1989|