The role of the carbohydrate moiety on the size heterogeneity and immunologic determinants of human testosterone-estradiol-binding globulin.
Human testosterone-estradiol-binding globulin (hTeBG) has been purified to apparent homogeneity by several laboratories using procedures which, in most instances, were labor intensive. In this report, hTeBG was purified from pregnancy serum by a newly developed two step procedure involving sequential affinity chromatography and ion-exchange high performance liquid chromatography (ion-exchange HPLC). The purity of the final product was confirmed by silver stained SDS-polyacrylamide gel and reverse phase HPLC monitored at 206 nm. hTeBG purified by ion-exchange-HPLC maintained binding activity by Dextran coated charcoal (DCC) assay and size heterogeneity on SDS-polyacrylamide gels which were indistinguishable from those of the proteins purified by conventional chromatography. Removal of the carbohydrate moiety from the molecule by both enzymatic and chemical treatment reduced the apparent molecular size and eliminated lectin binding of hTeBG subunits. Deglycosylation did not, however, abolish or alter the distribution of the protomeric forms of this subunit. We conclude that hTeBG is a dimer whose monomer exhibits two protomeric forms which is not a result of carbohydrate heterogeneity. In addition, disialylated and deglycosylated hTeBG exhibited antigenic determinants identical to the native protein.
Cheng
C Y
CY
Musto
N A
NA
Gunsalus
G L
GL
Bardin
C W
CW
D002241:Carbohydrates / Q000276:immunology*; D002845:Chromatography; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000276:immunology*; D005260:Female; D006801:Humans; D046911:Macromolecular Substances; D008970:Molecular Weight; D019158:N-Acetylneuraminic Acid; D011247:Pregnancy; D012738:Sex Hormone-Binding Globulin / Q000276:immunology* / Q000302:isolation & purification; D012794:Sialic Acids / Q000276:immunology
D002241:Carbohydrates; D000939:Epitopes; D046911:Macromolecular Substances; D012738:Sex Hormone-Binding Globulin; D012794:Sialic Acids; D019158:N-Acetylneuraminic Acid