Iodoamino acid distribution in thyroglobulin iodinated in vivo and in vitro

Abstract

The amount and the distribution of the iodoamino acids in thyroglobulin iodinated in vivo were compared to those obtained by iodination in vitro. Normal bovine thyroglobulin (54 atoms I per molecule) and virtually iodine-free human goiter thyroglobulin (0.5 atom I per molecule) were iodinated with 100 and 200 moles 131I 2 per molecule thyroglobulin at pH 9.0 and at pH 7.2, and then hydrolyzed with pronase. The distribution of the radioactivity among the various iodoamino acids, including 3,3′,5′-triiodothyronine, 3′,5′-diiodothyronine, and monoiodohistidine, in the pronase digest was determined by cation exchange chromatography and thin-layer chromatography. Native rat thyroglobulin, iodinated in vivo by equilibrium labeling, was analyzed in the same way. In equilibrium-labeled native thyroglobulin small amounts of 3,3′,5′-triiodothyronine, 3,3′- and 3′,5′-diiodothyronine, and monoiodohistidine were present in addition to mono- and diiodotyrosine, 3,3′,5-triiodothyronine, and thyroxine. In thyroglobulin which had been iodinated in vitro, considerably smaller amounts of the thyroid hormones thyroxine and 3,3′,5-triiodothyronine were present and the biologically inactive iodothyronines (3,3′,5′-triiodothyronine and 3′,5′-diiodothyronine) formed a proportionately higher percentage of the total iodothyronines. No labeled 3,3′-diiodothyronine could be detected in thyroglobulin iodinated in vitro. Monoiodohistidine, present in native thyroglobulin only in minute amounts, was found in considerably larger amounts after iodination in vitro. Thyroglobulin iodinated in vitro is less extensively hydrolyzed by pronase than native thyroglobulin which suggests structural differences between the two types of thyroglobulin. Although the various differences between thyroglobulin iodinated in vivo and in vitro may be partly due to species differences, they seem to arise mainly from differences in the mechanism of the two types of iodination.