Low Sialic Acid Content Research Articles

Further characterization of human thyroxine-binding globulin

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.

Isolation of desialylated and low-iodinated thyroglobulin in an experimental rat thyroid tumour.

ABSTRACT In an experimental rat thyroid tumour (Wollman line 1-1C2) thyroglobulin (TG) was isolated from soluble iodoproteins by ammonium sulphate fractionation and density gradient ultracentrifugation. The isolated TG was characterized by polyacrylamide gel electrophoresis, microimmunodiffusion, immunoelectrophoresis, analytical ultracentrifugation and by chemical determination of sialic acid and iodine content. Tumour thyroglobulin has a sedimentation coefficient of 17.7S, which is similar to that of enzymatically desialylated thyroglobulin (18.4S) in the normal rat; tumour TG shows a slower electrophoretic mobility than native TG and similar to rat desialylated TG. In double immunodiffusion tumour TG is shown to precipitate with an identity reaction with both native and desialylated rat TG; in immunoelectrophoresis it has a slower migration than native TG and is very close to that of enzymatically desialylated TG. Tumour TG has an iodine content of 0.02 % and a sialic content of 0.15 %. Chromatography of the 125I-labelled tumour TG revealed the presence of mono- and diiodothyrosines, a very small amount of triiodothyronine but no thyroxine. These results suggest that tumour TG, isolated in the experimental rat thyroid tumour, previously shown not to incorporate sialic acid into thyroglobulin, has a different electrophoretic mobility from native TG but is very similar to desialylated rat TG and has a very low sialic acid and iodine content as well as a striking decrease in thyroxine.

Evidence for an intrinsic gonadotropic activity of ovine LH in the lizard

Ovine LH that was highly purified by repeated precipitation in 0.5 saturated ammonium sulfate (to remove FSH) or treated with an antiserum that is highly effective in blocking FSH activity still exhibited both spermatogenic and androgenic stimulating activity in the hypophysectomized male lizard ( Anolis carolinensis). It is thus concluded that the ovine LH molecule does possess intrinsic gonadotropic activity in the lizard although it is considerably less potent than the FSH. Neuraminidase caused a marked (70%) reduction of the gonadotropic activity of LH in the lizard indicating that, contrary to previous reports, ovine LH probably does contain a small amount of sialic acid that is important for some of its biological activities. On the basis of the relatively low sialic acid content of this LH preparation (about 0.36 residues per molecule) it is proposed that the small fraction of molecules containing sialic acid contributes disportionately to the total gonadotropic activity of the ovine LH in the reptile. These findings suggest that the large disparity in relative potencies of mammalian FSH and LH in reptiles may be due partly to differences in their biological half-lives.

Studies on the Catabolism of γG Subunits and Chains

Summary The catabolism of I131 labeled H- and L-chain of RGG and HGG, Bence Jones protein and H-chain protein was studied in rabbits. Evidence is presented that the molecular structure responsible for the persistence in the circulation of γG is localized in the H-chain. The H-chain was slowly eliminated from the circulation similar to reduced and alkylated γG. The H-chain protein, obtained from patients with H-chain disease, was eliminated from the circulation with a half-life not significantly different from that of HGG. Two populations of H-chain protein with different electrophoretic mobilities were separated by DEAE-cellulose chromatography. Both populations were catabolized at the same rate. The rabbits did not excrete the H-chain protein in the urine. The L-chain and the Bence Jones protein were very rapidly eliminated from the circulation. A significant amount of the L-chain obtained from RGG but not from HGG was excreted in the urine as protein bound radioactivity. Sialic acid was shown not to be an important part of the structure necessary for the persistence in the circulation of γG. Removal of sialic acid did not change the rate of elimination of HGG, HGG papain Fc fragment and an H-chain protein preparation with a low sialic acid content. In contrast, a large portion of the H-chain protein preparations with a high sialic acid content was rapidly eliminated and catabolized after removal of sialic acid.

Neuraminidase in Mammalian Brain

CONTAMINATION of serum with neuraminidase-producing bacterial cells promptly produces changes in the electrophoretic pattern. The mobility (surface charge) of the α1-globulins (orosomucoid and α1-antitrypsin), of some α2-globulins (for example, haptoglobin) and of the β1-globulin, transferrin, decreases on release of sialic acid from the molecular surface1. Vulpe2 has published electrophoretic protein patterns of rabbit sera before and during development of experimental encephalomyelitis induced by injections of a brain suspension in combination with Freund's adjuvant. The changes in serum presented are similar to those produced by neuraminidase. Furthermore, the transferrin of human cerebrospinal fluid is electrophoretically heterogeneous3, indicating that part of it has lower sialic acid content than the most common transferrin type of Caucasians. These two observations prompted the present investigation of mammalian brain for neuraminidase. This enzyme is common among microorganisms, and has been found in chick-embryo chorioallantois but not in mammalian tissue4.