|Soffer, R. L.; Hechtman, P.; Savage, M.
|J Biol Chem
|An enzyme which catalyzes the transamination of L-triiodothyronine and of halogenated and nitrated tyrosines has been detected in the soluble fraction of a variety of rabbit organs. Kidney and liver had the highest specific activities of this enzyme which was isolated from the latter source. The most purified fraction was enriched approximately 1,900- and 650-fold for L-triiodothyronine and 3,5-dinitro-L-tyrosine aminotransferase activities, respectively.
Electrophoresis on polyacrylamide gels suggested that it was about 70% pure. The molecular weight estimated by gradient centrifugation was 95,000, and a value of 48,000 was obtained by disc gel electrophoresis in sodium dodecyl sulfate.
Activities for the transamination of 3-iodo-L-tyrosine, 3,5-diiodo-~tyrosine, and L-thyroxine were also copurified during the isolation procedure. The apparent Michaelis constants and turnover numbers for the different substrates were: L-triiodothyronine, 0.034 mu and 1.9 moles per min per mole of enzyme; L-thyroxine, 0.2 and 1.4; 3,5-diiodo-Ltyrosine, 0.9 and 66; 3-iodo-L-tyrosine, 3.8 and 221; 3,5-
dinitro-L-tyrosine, 4.4 and 2391.
Transamination of either L-triiodothyronine or 3, S-dinitro- L-tyrosine was dependent upon the presence of oc-ketoglutarate and of 2-mercaptoethanol. There was a partial requirement for pyridoxal phosphate. Triiodothyronine could be regenerated from its reaction product using purified aminotransferase and L-glutamate. When 3,5-dinitro-I,-tyrosine was used as substrate, molar equivalents of 3,5-dinitro-p-hydroxyphenylpyruvic acid and glutamic acid were produced.
The specificity of the enzyme for amino acids was markedly influenced by the cy-keto acid substrate. Pyruvate or oxalacetate could replace ar-ketoglutarate when L-triiodothyronine was the amino group donor, but were ineffective with 3,5-dinitro-L-tyrosine and only partially effective with the iodinated tyrosines. Thus, in the presence of these alpha-keto acids, the enzyme became relatively specific for L-triiodothyronine.