Properties of a peroxidase purified from beef thyroid tissues

Abstract

Abstract 1. 1. A peroxidase was purified from beef thyroid tissues by column chromatography on ion-exchange CM-Sephadex. Some of the properties of the purified enzyme were studied. 2. 2. By means of electrophoresis on polyacetate strips the purified enzyme was found to consist of one protein component. This single component demonstrated peroxidase activity when stained with o-dianisidine in the presence of H2O2. 3. 3. The molecular weight of the beef thyroid peroxidase was estimated to be approx. 50 000 by sucrose gradient centrifugation using as a reference a purified preparation of dog myeloperoxidase. 4. 4. Protoporphyrin IX was identified as the prosthetic group by spectral analysis of the acid methylethyl ketone extract and the pyridine haemochrome of the enzyme. 5. 5. Spectral properties of the enzyme complexes with CN- and CO, as well as its reduction by sodium dithionite, were distinctively different from those of bovine haemoglobin. 6. 6. Kinetic studies using o-dianisidine as a hydrogen donor showed that the Km of the enzyme for H2O2 was between 1.5 to 2.0 · 10−5 M. 7. 7. The peroxidase activity of the enzyme was inhibited by SCN-, CN-, N3-, F-, I-, propylthiouracil and Tapazole but not by p-hydroxymercuribenzoate or ClO4- as shown by kinetic studies. Thyroid-stimulating hormone did not show any effect. 8. 8. The enzyme catalyzed also the iodination of tyrosine actively in the presence of added H2O2 or a H2O2-generating system. The iodination of tyrosine by this enzyme occurred optimally at pH 6. Thyroglobulin was iodinated much less actively. 9. 9. The enzymic iodination of tyrosine was inhibited by the inhibitors mentioned above, except iodide, and was not affected by thyroid-stimulating hormone.