Characteristics of Iodothyronine Tyrosyl Ring Deiodination by Rat Cerebral Cortical Microsomes*

Abstract

We studied the biochemical properties of tyrosyl ring (5-) deiodination of L-T₃ and L-T₄ by cerebrocortical microsomes from euthyroid rats. Incubations contained radioiodinated L-T₃) or L-T₄ and dithiothreitol (DTT), and products were analyzed by paper chromatography. The pH maximum was 7.5 for 5-deiodination of both T₃ and T₄. Increasing the DTT concentration from 5 to 200 mM caused a progressive increase in the 5- deiodination rate of 2 nM T₃, but at 500 mM DTT the rate decreased. When three DTT concentrations (5.25, 10.1, and 20.1 mM) were used, double reciprocal plots of 5-deiodination rate as a function of T₃ concentration (from 0.3–40 nM) showed a sequential type kinetic pattern, with a limiting Kn of 1.7 nM L-T₃. Cerebral microsomes from 4-day-old rats had an apparent K_m for L-T₃ similar to that of cerebrocortical microsomes from adult rats, but the neonatal tissues had a 3.5- to 10-fold higher apparent maximum velocity. At 50 mM DTT, L-T₃ and L-T₄ each inhibited 5-deiodination of the other. The apparent K_m and K_i for L-T₃ were 5.5 and 8.0 nM, respectively, the apparent K_m and K_i for L-T₄ were 37 and 48 nM, respectively, and the apparent maximum velocities for 5-deiodination of T₃ and T₄, respectively, were 134 and 144 fmol/min/mg protein. There was dose-dependent inhibition of L-T₃ 5-deiodination by tetraiodothyroacetic acid, D-T₃, tetraiodothyroacetic acid, 3,5-L-diiodothyronine, 3,3′-L-diiodothyronine, iopanoic acid, rose bengal, L-rT₃, erythrosine, bromphenol blue, and anilinonaphthalenesulfonic acid (in decreasing order of potency). There was no effect on reaction rate by addition of 3′-L-monoiodothyronine, L-thyronine, DIT, amiodarone, dicoumarol, Nal, propylthiouracil (PTU), sodium salicylate, sodium fluoride, EDTA, CaCl₂, or MgCl₂. These data demonstrate that the properties of the rat brain iodothyronine tyrosyl ring deiodinase differ from those of both the PTU-sensitive and PTU-insensitive pathways of 5′-deiodination in rat brain and also differ from the properties of the iodothyronine 5-deiodinase in rat liver.