Studies on the Role of Glycosylation for Human Corticosteroid-Binding Globulin: Comparison with That for Thyroxine-Binding Globulin*

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Abstract
The role of glycosylation on the secretion and the stability of human corticosteroid binding globulin (CBG) was studied. Cells of the human hepatoma line were labeled by [35S]methionine in presence of or absence of tunicamycin (TM). Media or cells were harvested at 0, 3, 6, and 20 h after the addition of excess unlabeled methionine. Media and cell lysates were incubated with anti-CBG serum and immune complexes were precipitated with Staphylococcus aureus protein A (Pansorbin). Immunoprecipitates were analyzed by fluorography after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Immunoprecipitation of T4-binding globulin (TBG) was also carried out with anti-TBG serum. Fluorographic analysis revealed three forms of CBG: CBG1, a glycosylated, mature, and secretory form with apparent mol wt of 70 K; CBG2, a glycosylated precursor which due to incomplete carbohydrate processing has an apparent mol wt of 54 K; and CBG3 a nonglycosylated form consisting of the 40 K core protein. In absence of TM, CBG1 was observed in media and CBG2 was detected in cell lysates. The proportion of CBG1 increased during the chase, whereas that of CBG2 decreased, indicating that CBG was secreted after processing of the oligosaccharides on CBG2. In presence of TM, CBG3 was found both in media and cell lysates. The sum of CBG3 in the medium and the cell lysate decreased during the chase, whereas that of CBG1 and CBG2 remained unchanged. Similar to CBG, TBG1 (mature form, 60 K) and TBG2 (partially processed glycosylated form, 54 K) were observed in media and cell lysates, respectively, in absence of TM. However, TBG3 (nonglycosylated, 44 K) was not detected in medium. These results indicate that glycosylation is not a key factor for the secretion of CBG but is important for its stability. On the other hand the glycosylation is indispensable for the secretion of TBG.

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