The goal of the project is to increase our understanding of the structure-function relationships of oligosaccharide moieties on glycoprotein hormones and related molecules. The molecules under investigation are human chorionic gonadotropin (hCG), free alpha molecules associated with pregnancy or malignancy, hCG-beta subunit, beta-core fragment and corticosteroid binding globulin (CBG). Carbohydrate modifications, resulting in a variety of branched oligosaccharide structures, occur on all glycoproteins prior to secretion. These modifications can effect virtually every aspect of the molecule's behavior, including receptor binding and signal transduction, yet the underlying regulatory mechanisms remain unclear. We have investigated changes in the oligosaccharide moieties as a function of the following parameters: 1. Synthetic environment. Interaction of subunits can either promote or restrict modifications on the oligosaccharide structures produced in the final secreted protein. When hCG-beta subunit combines with hCG-alpha subunit prior to carbohydrate processing, hCG-beta prevents addition of some branches to hCG-alpha whereas the uncombined (free) alpha molecules receive the additional branches. The presence of these branched structures on free alpha prevent it from subsequently combining with beta to form intact hormone. 2. Gestational development. Glycosylation changes as a function of gestational age. Molecules produced in late pregnancy are more highly branched than those of early pregnancy. 3. Cellular differentiation. Oligosaccharide changes occur in placental cytotrophoblasts as they begin to undergo cellular differentiation. 4. Species specificity. We showed that rat CBG contained a carbohydrate composition strikingly different from that of human CBG. Variations in glycosylation may account for differences in receptor-hormone interactions observed between species. 5. Cellular catabolism. Beta-core fragment is derived from hCG-beta subunit and is the most abundant hCG-related component found in pregnancy urine. Beta-core has important potential as a clinical marker for a number of pathologic conditions, including a variety of malignancies. We have characterized the carbohydrate structures on beta-core and compared them to HCG-beta. The structural analyses described herein further our knowledge about the complexities associated with glycosylation. We plan to probe how the differences observed in glycosylation effect the bioactivity of these molecules.
