Comparison of kidney and brown adipose tissue iodothyronine 5'-deiodinases

We have examined the influence of assay conditions on the 6-n-propyl-2-thiouracil (PTU) sensitivity of the iodothyronine 5'-deiodinase in brown adipose tissue (BAT) from hypothyroid rats. These results were compared with similar studies of 5'-deiodinase activity in kidney microsomes from euthyroid animals. Even though BAT microsomes contain largely type II (PTU-insensitive) deiodinase activity, the 5'-deiodination of T4 can be inhibited by PTU if the dithiothreitol (DTT) concentration in the assay is reduced to 5 mM or less. The apparent Ki for PTU of BAT microsomes was 4.3 mM at 5.0 mM DTT and 0.41 mM at 0.5 mM DTT. The kinetics of inhibition were noncompetitive. With kidney microsomes, PTU inhibition of rT3 5'-deiodination was both time and enzyme/substrate ratio dependent. For example, using 1 microgram microsomal protein, 2 nM rT3, and 5 mM DTT, the inhibitory effect of PTU was not maximal until 12 min after PTU addition. At stable reaction velocities PTU inhibition was uncompetitive, and the Ki was about 1 microM. Deiodination by kidney microsomes was completely inhibited by 50 microM PTU. Even though it is possible to inhibit the type II 5'-deiodinase activity with high concentrations of PTU (in the presence of low DTT concentrations), the deiodinase in kidney is about 1000-fold more sensitive to PTU. By these criteria the kidney microsome 5'-deiodinase is type I.