Ptocess of Iodination of Thyroglobulin and Its Maturation

II. Properties and Distribution of Thyroglobulin Labeled In Vitro or In Vivo with Radioiodine, ³H-Tyrosine, or ³H-Galactose in Rat Thyroid Glands

Abstract

With the aim of obtaining information on the process of iodination of thyroglobulin, the properties and subcellular distribution of thyroglobulin labeled with radioiodine, ³H-tyrosine, or ³H-galactose were studied. The following results were obtained for 17-19S thyroglobulin isolated from rat thyroid lobes labeled in vitro. (a) The effect of sodium dodecyl sulfate (SDS) concentration (0.1-2.0 mM) on the dissociability of the proteins into 12S subunits showed that ³H-labeled, ¹³¹I-labeled, and preformed thyroglobulin behaved very differently; their dis-sociability decreased in that order. In addition, 0.3 mM SDS is most suitable for discriminating among these species. (b) The amount of 0.3mm SDS-resistant ¹³¹I-thyroglobulin increased with the time of incubation of the lobes or with the amount of iodine atoms incorporated by chemical iodination. (c) Digestion of ³H-tyrosine-labeled thyroglobulin showed that ³H-monoiodotyrosine and ³H-diiodotyrosine were present after incubation of the lobes for 180 min. (d) The dissociability of ³H-galactose-labeled 17-19S thyroglobulin was higher than that of ¹³¹1I-labeled protein, but its elution pattern on DEAE-cellulose chromatography resembled that of the latter. (e) ¹³¹I-Thyroglobulin was scarcely found in the incubation medium, although a considerable amount of 19S thyroglobulin was released into the medium during the incubation. As for the lobes, a significant amount of ¹³¹I-radioactivity as well as ³H-radioactivity was found in cytoplasmic particulates, especially in fractions containing apical vesicles and rough microsomes. On the other hand, the following results were obtained for 17-19S thyroglobulin isolated from rats injected with ¹²⁵I. (a) Dissociability of the protein by 0.3 mM SDS and analysis of ¹²⁵I-iodoarnino acids of pronase digests showed that the iodina-tion process was essentially similar to the case of in vitro incorporation, but was faster. (b) The effect of cycloheximide treatment showed that the relative reduction of 0.3 mM SDS-