Expression and differential glycosylation of human sex hormone-binding globulin by mammalian cell lines.
The human sex hormone-binding globulin (SHBG) gene is responsible for the production of plasma SHBG by the liver and androgen-binding protein in the testis. Cell-specific glycosylation events during synthesis may account for minor differences in the biochemical properties of SHBG and androgen-binding protein, and we have, therefore, expressed a human SHBG cDNA in chinese hamster ovary (CHO) cells and a mouse hepatoma cell line (BW-1), and compared the products to SHBG in serum. The SHBG produced in this way is a homodimer of subunits that exhibit size microheterogeneity similar to SHBG in human serum, and its affinity for 5 alpha-dihydrotestosterone (Kd = 0.6 nM) and other steroids is essentially identical to that of natural SHBG. When medium from transfected CHO and BW-1 cells was subjected to Concanavalin-A (Con-A) chromatography, the relative amounts of SHBG retained by Con-A were 74% and 86%, respectively. In addition, when SHBG produced by CHO cells was separated into two fractions by Con-A chromatography and analyzed by polyacrylamide gel electrophoresis, SHBG that did not interact with Con-A migrated with a slightly larger apparent mol wt than that of SHBG that binds Con-A; this can be explained by the presence of triantennary, rather than biantennary, N-linked oligosaccharide chains. These data also demonstrate that the subunit microheterogeneity associated with plasma SHBG reflects differences in glycosylation during synthesis, which appear to be cell type specific.
Bocchinfuso
W P
WP
Department of Obstetrics and Gynecology, University of Western Ontario, London, Canada.
Warmels-Rodenhiser
S
S
Hammond
G L
GL
D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D016466:CHO Cells; D002460:Cell Line; D002846:Chromatography, Affinity; D003208:Concanavalin A; D006224:Cricetinae; D013196:Dihydrotestosterone / Q000378:metabolism; D000939:Epitopes / Q000032:analysis; D015870:Gene Expression; D005822:Genetic Vectors; D006031:Glycosylation; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D051379:Mice; D011499:Protein Processing, Post-Translational; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D012738:Sex Hormone-Binding Globulin / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D014162:Transfection
D000939:Epitopes; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D012738:Sex Hormone-Binding Globulin; D013196:Dihydrotestosterone; D003208:Concanavalin A; D012313:RNA