Further characterization of human thyroxine-binding globulin

Korcek, Ladislav; Tabachnick, Milton

doi:10.1016/0005-2795(74)90029-4

Cited by 32 publications

(4 citation statements)

References 27 publications

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“…3 . Moist Epoxy-activated Sepharose 6B (45 ml; Pharmacia) was then added to the T 3 solution, and the gel suspension was gently shaken overnight at room temperature.…”

Section: Methodsmentioning

confidence: 99%

Thyroxine-Binding Globulin: Investigation of Microheterogeneity*

Gärtner

Henze

Horn

et al. 1981

The Journal of Clinical Endocrinology & Metabolism

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“…3 . Moist Epoxy-activated Sepharose 6B (45 ml; Pharmacia) was then added to the T 3 solution, and the gel suspension was gently shaken overnight at room temperature.…”

Section: Methodsmentioning

confidence: 99%

Thyroxine-Binding Globulin: Investigation of Microheterogeneity*

Gärtner

Henze

Horn

et al. 1981

The Journal of Clinical Endocrinology & Metabolism

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“…Upon photolysis, one band was strongly labeled (lane d). This band, which runs just ahead of the overloaded serum albumin band, is appropriate for the reported size estimates of TBG (Nilsson & Peterson, 1975;Gershengom et al, 1977; Korcek & Tabachnick, 1974). A light background due to labeling of serum albumin was visible in this lane as well.…”

Section: Resultsmentioning

confidence: 60%

Photoaffinity labeling of human thyroxine-binding prealbumin with thyroxine and N-(ethyl-2-diazomalonyl)thyroxine

Somack¹,

Nordeen²,

Eberhardt³

1982

Biochemistry

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To facilitate studies of thyroid hormone-binding proteins, we have synthesized and tested the photoaffinity analogues N-(ethyl-2-diazomalonyl)-3,5,3'-triiodothyronine (EDM-T3) and N-(ethyl-2-diazomalonyl)thyroxine (EDM-T4). The binding affinities of L-EDM-T4 and D-EDM-T4 to human thyroxine-binding prealbumin were 4% and 13.2%, respectively, that of L-thyroxine (L-T4). For comparison the affinities of L-EDM-T3 and D-EDM-T3 to crude rate liver nuclear receptor preparation were 0.1% and 0.7%, respectively, that of L-triiodothyronine (L-T3). Photolysis of prealbumin-[125I]-L-EDM-T4 complexes at 254 nm resulted in covalent linkage of [125I]-L-EDM-T4 to prealbumin as judged by sodium dodecyl sulfate gel electrophoresis. Virtually no labeling was observed in the absence of photolysis. Photolabeling of prealbumin was specific for the high-affinity hormone binding site since it was (a) completely blocked during photolysis in the presence of excess 3,5,3',5'-tetraiodothyroacetic acid, (b) saturated at high [125I]-L-EDM-T4 concentrations, (c) prevented when the hormone binding site had been previously blocked by dansylation of prealbumin, and (d) blocked competitively by T3 and T4 with inhibition constants (K1) similar to the dissociation constants (Kd) for these ligands. Analysis of prealbumin photolabeling by direct attachment of [125I]-L-EDM-T4 or attachment of unlabeled L-EDM-T4 followed by titration of the remaining sites with [125I]-L-T4 indicated a photolabeling efficiency of 54-61% at 63-67% site occupancy. After destruction of the diazo group by preirradiation, L-EDM-T4 was found to label prealbumin after further irradiation; 19-26% photolabeling efficiency could be achieved by using preirradiated reagent at 87-91% site occupancy. This carbene-independent photoattachment was also specific for a high-affinity hormone binding site. The mechanism of the carbene-independent process may involve attachment via radical formation following photoinduced loss of the thyronine ring iodine. Accordingly, specific covalent cross-linking of L-T4 to prealbumin was demonstrated; however, the photolabeling efficiency was much lower than that with preirradiated EDM-T4. Both EDM-T4 and T4 were employed to photolabel prealbumin, thyroxine binding globulin, and albumin in unfractionated human serum.

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“…Thyroxine-binding globulin, the major plasma transport protein for thyroid hormones, has been purified from human blood [1][2][3]. Several groups have studied its chemical properties especially as they relate to its interaction with thyroid hormones [4][5][6][7][8][9] and metabolic clearance rates have been reported recently in différent clinical states [10,11].…”

Section: Introductionmentioning

confidence: 99%

Thyroxine-binding globulin biosynthesis in isolated monkey hepatocytes

Glinoer

Gershengorn

Robbins

1976

Biochimica et Biophysica Acta (BBA) - Nucleic Acids and Protein

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Thyroxine-binding globulin biosynthesis was demonstrated in hepatocytes isolated from normal adult Rhesus monkeys. Dispersed cells were obtained by in situ liver perfusion with collagenase, hyaluronidase and EDTA. Conditions for optimum cell survival and incorporation of radioactive leucine into newly synthesized proteins were defined. Protein synthesis, and specifically thyroxine-binding globulin synthesis, were shown to continue throughout the incubation period, while cell survival remained high (75% excluded trypan blue after 6h). Incubation medium, cytosol and a particulate fraction (extracted with digitonin) were analyzed for thyroxine-binding globulin. After extensive dialysis and purification by affinity chromatography, newly synthesized thyroxine-binding globulin was identified by specific double-antibody immunoprecipitation and by immunodiffusion and immunoelectrophoresis with autoradiography. Newly synthesized thyroxine-binding globulin was present after 4 h of incubation. After 6 h, the total synthesized had increased to 150% of the 4 h value, while the fraction present in the medium and increased to 300%, indicating probable thyroxine-binding globulin secretion

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Further characterization of human thyroxine-binding globulin

Cited by 32 publications

References 27 publications

Thyroxine-Binding Globulin: Investigation of Microheterogeneity*

Thyroxine-Binding Globulin: Investigation of Microheterogeneity*

Photoaffinity labeling of human thyroxine-binding prealbumin with thyroxine and N-(ethyl-2-diazomalonyl)thyroxine

Thyroxine-binding globulin biosynthesis in isolated monkey hepatocytes

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