The goal of the project is to increase our understanding of the structure- function relationships and glycobiology of the oligosaccharide moieties on glycoprotein hormones. The molecules currently under investigation are, human chorionic gonadotropin (hCG), gonadotropin free alpha subunits associated with pregnancy or malignancy, and pituitary free alpha subunits secreted throughout the normal menstrual cycle. We have shown that the free alpha subunit purified from pregnancy urine stimulates secretion of prolactin from primary cultures of human decidual cells in a dose- dependent manner. These findings indicate that free alpha may be a glycoprotein hormone with a function that is independent of hCG. Carbohydrate modifications, resulting in a variety of branched oligosaccharide structures, occur on all glycoproteins prior to secretion. These modifications can affect virtually every aspect of the molecule's behavior, including receptor binding and signal transduction, yet the underlying regulatory mechanisms remain elusive. We have investigated functional aspects of the oligosaccharide moieties on free alpha, demonstrating that specific types of carbohydrate modifications on free alpha can either prevent or facilitate combination with hCG-beta to form intact hormone. We have found that the free alpha subunits that are located in fetal compartments in high concentrations during early pregnancy exhibit physical properties that are similar to those of the free alpha subunit isolated from pregnancy urine. It is likely that oligosaccharide moieties on this early pregnancy free alpha function to maintain the molecule in an uncombined form. We have identified differences in the oligosaccharide moieties of free alpha and hCG as a function of gestational development; in particular, the glycosylation of free alpha changes dramatically as a function of gestational age. Molecules produced in late pregnancy are more highly branched and are much more extensively fucosylated than those of early pregnancy. We plan to examine how the differences observed in glycosylation affect the bioactivity of these molecules.
