Further characterization of human thyroxine-binding globulin

Abstract

Thyroxine-binding globulin has been purified from human blood plasma by a three-step procedure consisting of affinity chromatography on thyroxine-Sepharose, chromatography on DEAE-Sephadex A-50, and preparative electrophoresis in polyacrylamide gel. By ultracentrifugal analysis the molecular weight of thyroxine-binding globulin was found to be 60 700 compared to an estimated value of 65 000 by sodium dodecylsulfate-polyacrylamide gel electrophoresis. Thyroxine-binding globulin contained about 13% carbohydrate which included per mole of protein, 2 residues glucose, 6 residues galactose, 12 residues mannose, 12 residues glucosamine, and 6 residues sialic acid. Analysis by gas-liquid chromatography revealed the presence of long-chain fatty acids associated with thyroxine-binding globulin. A second form of thyroxine-binding globulin which moved appreciably slower electrophoretically than thyroxine-binding globulin was also isolated. The slower moving thyroxine-binding globulin had a low sialic acid content, 0.6 residue per mole, and is apparently produced in significant quantity by desialylation of thyroxine-binding globulin during the purification procedure. Slow thyroxine-binding globulin had a molecular weight in the vicinity of 57 500 as determined by sodium dodecylsulfate-polyacrylamide gel electrophoresis. No evidence was found for a subunit structure in either thyroxine-binding globulin or the slower moving form of thyroxine-binding globulin.