V M Nikodem — Page 4 | PubMed Health

PubMed1981

Two-dimensional gel analysis of rat liver nuclear proteins after thyroidectomy and thyroid hormone treatment.

The composition of nucleoproteins in normal rat liver, in livers from thyroidectomized animals, and in thyroidectomized animals treated with 3,3',5-triiodo-L-thyronine was examined by two-dimensional gel electrophoresis. A computer program was designed to measure the relative concentrations of each protein. Approximately 500 protein subunits could be distinguished. Of these, 102 either disappeared or were markedly decreased after thyroidectomy. Shortly after administration of the thyronine (6 hr), 13 proteins reappeared and, by 24 hr, 67 additional proteins had reappeared. These changes in protein concentrations are both the greatest in number and have the largest reported as an effect of thyroid hormone. Twenty-four hr after triiodothyronine administration, some 22 proteins seen in normal liver could not be found. Two new proteins appeared in thyroidectomized rat liver nuclei. We conclude that a major and rapid effect of thyroid hormone is exerted on hepatic nucleoproteins.

Animals↗

PubMed1980

Affinity labeling of rat liver thyroid hormone nuclear receptor.

The thyroid hormone receptor from rat liver nuclei has been covalently labeled with the N-bromoacetyl derivatives of L-thyroxine (T4) and 3,3',5-triiodo-L-thyronine (T3). Displacement binding studies showed that, in the presence of 100-fold molar excess of unlabeled N-bromoacetyl-T3 or T4, binding of [125I]T3 or [125I]T4 was nearly totally inhibited. Heat inactivation of the receptor (55 degrees C for 15 min) resulted in parallel losses in the binding of T3 (95%) and N-bromoacetyl-T3 (93%). These results indicated that T3 and T4 and their bromoacetyl derivatives compete for the same binding site. The nuclear receptor showed identical behavior in high-pressure liquid chromatography (HPLC) whether bound to T3 or T4 or covalently labeled with their bromoacetyl derivatives. HPLC provided a single-step 100-fold purification of the nuclear receptor. Na-DodSO4 gel electrophoresis of the nuclear receptor labeled with N-bromoacetyl derivatives of [125I]T3 or [125I]T4 showed one major radioactive component with a molecular weight of 56,000. Furthermore, in the absence of denaturant, the nuclear receptor either bound to [125I]T3 or covalently labeled with N-bromoacetyl-[125I]T3 showed identical mobility. These results suggested that the nuclear receptor is a single polypeptide chain and binds either T3 or T4. Nuclear receptors covalently linked with N-bromoacetyl derivatives of [125I]T3 or [125I]T4 may be useful as a marker for the preparative purification of receptor.

Affinity Labels↗