3′5′-Diiodothyronine to 3′-Monoiodothyronine Conversion in the Fed and Fasted Rat: Enzyme Characteristics and Evidence for Two Distinct 5′-Deiodinases*

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Abstract
The present study investigated the 5′-monodeiodination of 3′5′-diiodothyronine (3′,5′T2) to 3′-monoiodothyronine (3′T1), a reaction involving deiodination at the same position as is needed for T4 to T3 and for rT3 to 3,3′-diiodothyronine (3,3′T2) conversion. 3′5′T2 to 3′T1 conversion appears to be enzymatic in nature, being dependent on temperature and pH. Consistent with previous observations for T4 and rT3, 3′5′T2 5′-deiodination occurred primarily in the liver and kidney, but, unlike these other iodothyronines, 3′5′T2 to 3′T1 conversion was active in cytosol as well as in microsomes. Characteristics of the microsomal enzymatic activity included heat inactivation (56 C), a broad pH plateau (pH 4.9–7.6), and an apparent Km of 8.6 × 10-9 M. Cytosolic activity occurred with a narrower pH peak (pH 5–6.5), and had a different apparent Km of 2.4 × 10-7 M. The activity of both subcellular enzymes was inhibited by iopanoic acid and propylthiouracil. Dithiothreitol, while stimulating a 5-to 7-fold increase in microsomal 3′5′T2 5′-deiodination, produced less than a 2-fold increase in cytosol activity. Fasting, a condition known to inhibit both T4 and rT3 5′-deiodination, produced a 68% decrease (1.16 ± 0.29 vs. 3.61 ± 0.97 ng/mg protein; mean ± SE) in hepatic T4 to T3 conversion, yet had no effect on 3′,5′T2 to 3′T1 conversion in rat liver or kidney homogenates. Cellular fractionation studies indicated that liver microsomal 3′,5′T2 deiodination was diminished by 46% (71.6 ± 19.8 vs. 131.6 ± 14.4 pg/mg protein; P < 0.025) after a 3-day fast, whereas cytosol activity was unchanged (23.7 ± 10.2 vs. 22.8 ± 4.9 pg/mg protein). Treatment with T4 (0.8 μg/100 g BW·day) failed to correct the reduced 3′,5′T2 5′-deiodinase activity in microsomes from fasted rats [79.7 ± 17.4 (fasted) vs. 311.9 ± 33.8 (control) pg/mg protein P < 0.001]. However, the addition of 5 mM dithiothreitol to the incubation medium increased hepatic microsomal 5′-deiodinase activity for 3′5′T2 7- and 12-fold in the fed and fasted animals, respectively. These results indicate that: 1) 3′5′T2 to 3′T1 conversion proceeds enzymatically in liver and kidney, 2) enzyme activities having different kinetics exist in both microsomes and cytosol, 3) fasting inhibits 3′5′T2 deiodination in microsomes but not in homogenates or cytosol, and 4) microsomal activity is restored by a sulfhydryl group-reducing agent, but not by physiological doses of T4. The data strongly suggest the existence of more than one 3′5′T2 5′-deiodinase in rat liver and kidney.