title: Preparation and properties of thyroxine-binding alpha globulin (TBG).
abstract: Thyroxine-binding alpha globulin (TBG) in human serum was isolated from Cohn fractions IV-5,6 and IV-4 by (1) chromatography on carboxymethyl (CM) cellulose, (2) gel filtration on Sephadex G-200, (3) chromatography on diethylaminoethyl-Sephadex, (4) a novel procedure of "double-gel" electrophoresis, and (5) preparative polyacrylamide gel electrophoresis. The protein was homogeneous by analytical disc gel electrophoresis, immunoelectrophoresis, and ultracentrifugal analyses (sedimentation velocity and sedimentation equilibrium), and after addition of thyroxine-(125)I showed a constant specific radioactivity on polyacrylamide electrophoresis. The sedimentation and diffusion coefficients were s(20, w), 3.0 x 10(-13) sec, and D(20, w), 8.05 x 10(-7) cm(2).sec(-1), and the molecular weight obtained by sedimentation equilibrium was 36,500. Gel filtration studies on Sephadex G-200 demonstrated that the protein had the same elution volume as that of native TBG in serum, apparently excluding the possibility of a subunit of the native protein. Chemical composition was ascertained by amino acid and carbohydrate analyses. The maximal thyroxine (T4)-binding capacity measured by reverse flow paper electrophoresis was 15,000 mug per g of protein, representing more than 2100 times that of the starting material, or about 5000 times that of whole serum. Based on the molecular weight obtained, the TBG preparation could bind 0.7 mole T4 per mole of protein, suggesting a single binding site. The association constant for T4 was estimated to be of the order of 10(10) by competitive binding studies employing TBG and T4-binding prealbumin (TBPA).
mesh_terms: D000510:Alpha-Globulins / Q000032:analysis / Q000302:isolation & purification*; D000596:Amino Acids / Q000032:analysis; D001665:Binding Sites; D001797:Blood Protein Electrophoresis; D002241:Carbohydrates / Q000032:analysis; D055598:Chemical Phenomena; D002621:Chemistry; D002845:Chromatography; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D004589:Electrophoresis, Disc; D004590:Electrophoresis, Paper; D006801:Humans; D007122:Immunoelectrophoresis; D007456:Iodine Isotopes; D008970:Molecular Weight; D011485:Protein Binding; D012709:Serum Albumin; D013974:Thyroxine; D014461:Ultracentrifugation
chemical_list: D000510:Alpha-Globulins; D000596:Amino Acids; D002241:Carbohydrates; D007456:Iodine Isotopes; D012709:Serum Albumin; D013974:Thyroxine