Studies of the in vitro and in vivo effects of conjugated steroids and carboxylic acids on hepatic tyrosine transaminase in the rat

Abstract

1. 1. Tyrosine transaminase (-tyrosine:2-oxoglutarate aminotransferase, EC 2.6.1.5) activity was lost rapidly in fresh rat-liver homogenates (pH 6.9), that were incubated at 38[deg]. The inactivation was paralled by the loss of the coenzyme but was not reversed by the subsequent addition of pyridoxal 5-phosphate.2. 2. The coenzyme, the keto acid substrates, and their anionic analogs retarded the inactivation and dissociation. Various anionic steroids and diethylstilbestrol disulphate (5[middle dot]10-4-5[middle dot]10-5 M) also retarded the inactivation and dissociation; free steroids were ineffective at saturation levels. Aromatic carboxylic acids were effective at 10-2-10-3 M, 5-hydroxytryptophan at 10-3 M, and -glutamate, bicarbonate, and Pi at 10-2 M. Several other amino acids and NaCl were ineffective at 10-2 M. Many of the in vitro stabilizing agents caused elevated levels of hepatic tyrosine transaminase when injected into adrenalectomized rats. In general, the most potent stabilizers were also the most effective agents in causing the elevated enzyme levels in vivo.3. 3. Estradiol disulfate and diethylstilbestrol disulfate also retarded the inactivation and dissociation that occured when the homogenates were incubated at 25[deg] in 1.1 or 2.2. M urea or when partially-purified tyrosine transaminase was incubated with trypsin (EC 3.4.4.4) or chymotrypsin (EC 3.4.4.5). The rate of inactivation in homogenates was not significantly changed by the presence of 0.001 M EDTA or mercaptoethanol nor by incubation with alkaline phosphatase (EC 3.1.3.1).4. 4. A small but significantly greater degree of association of the tyrosine transaminase with its coenzyme was found in rat-liver homogenates prepared 1 h after cortisol administration than in the injected controls that were sacrificed immediately. There was also a significantly slower rate of coenzyme dissociation in the 1-h animals. Similar doses of cortisone were ineffective in the latter case.