Human thyroid cyclic nucleotide phosphodiesterase: Its characterization and the effect of several hormones on the activity

Abstract

Cyclic AMP and cyclic GMP phosphodiesterase activities (3′5′-cyclic AMP 5′nucleotidohydrolase, EC 3.1.4.17) were investigated in the human thyroid gland from patients with hyperthyroidism. Low substrate concentration (0.4 μM) was used. About 60% of the cyclic-AMP and 80% of the cyclic-GMP hydrolytic activities in the homogenate were obtained in the soluble fraction ( 105 000 × g supernatant). The thyroid gland contains two forms of cyclic-AMP phosphodiesterase, one with a K m of 1.3 · 10 −5 M and the second with a K m 2 · 10 −6 M. Cyclic-AMP and cyclic-GMP phosphodiesterase were purified by gel filtration on a Sepharose-6B column. Cyclic-AMP phosphodiesterase activities were found in a broad area corresponding to molecular weights ranging from approx. 200 000 to 250 000 and cyclic-GMP phosphodiesterase activity was found in a single area corresponding to a molecular weight of 260 000. Cyclic-AMP phosphodiesterase activities were stimulated by the protein activator which was found in human thyroid and this stimulation was dependent on Ca 2+. Stimulation of cyclic-AMP phosphodiesterase by the activator was not significant even in the presence of enough Ca 2+. The effect of d,l-triiodothyronine, d,l-thyroxine, l-diiodotyrosine, l-monoiodotyrosine, l-thyronine, l-diiodothyronine, thyrotropin, hydrocortisone, adrenocorticotropin, cyclic-AMP and cyclic-GMP on the phosphodiesterase activities was studied. Cyclic-AMP, cyclic-GMP, d,l-triiosothyronine, d,l-thyroxine, adrenocorticotropin and hydrocortisone were found to inhibit the phophodiesterase. Triiodothyronine and thyroxine inhibited cyclic-AMP phosphodiesterase more effectively than cyclic-GMP phosphodiesterase. Thyroxine was a more potent inhibitor than triiodothyronine. The concentration of cyclic AMP producing a 50% inhibition of clclic-GMP phosphodiesterase activity was 5 · 10 −5 M, while the concentration of cyclic GMP producing a 50% inhibition of cyclic-AMP phosphodiesterase was 3 · 10 −3 M. Both cyclic-AMP and cyclic-GMP phosphodiesterase activities in the homogenate of hyperthyroidism, thyroid carcinoma and adenoma were higher than in normal thyroid tissue, when assayed with a low concentration of the substrate (0.4 μM). When a higher concentration (1 mM) of cyclic nucleotides was used as the substrate, cyclic-AMP hydrolytic activity in adenoma tissue was similar to that of normal tissue, while the other activities were higher than normal.