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A Study of Hepatic Low K_m Iodothyronine 5' -Monodeiodinase^*

RUBEN J. BOADO and INDER J. CHOPRA

Department of Medicine, University of California School of Medicine Los Angeles, California 90024

Abstract

To document the presence of a low K_m rT₃ and T₄-5' -monodeiodinase (5'MDL; K_m in nanomolar concentrations) in the liver and to study its characteristics in comparison with the high K_m 5'MD (5'MDH; K_m in micromolar concentrations), we incubated rat liver microsomal protein (20 µg for rT₃ substrate and 200 µg for T₄ substrate) with ¹²⁵I-labeled rT₃ or T₄ and dithiothreitol (DTT; up to 20 mM) for 5 min (for rT₃) or 30-120 min (for T₄) and determined the amount of ¹²⁵I liberated during incubation. Pilot studies had shown that the activity of rT₃ 5'MDH is markedly (85%) inhibited in the presence of 2 M NaCl, while the rT₃ 5'MDL is essentially unaffected, and both low and high K_m T₄ 5'MD are minimally (20%) inhibited. The representative kinetics of various substrates studied were: K_m, 13 nM for rT₃ 5'MDL, 640 for rT₃ 5'MDH, 26 for T₄ 5'MDL, and 3620 for T₄ 5'MDH; maximum velocity, 0.28 nmol/hmg protein for rT₃ 5'MDL, 46 for rT₃ 5'MDH, 0.002 for T₄ 5'MDL, and 0.46 for T₄ 5'MDH. Propylthiouracil and iopanoate inhibited all enzymic activities studied. The relative K_i values (micromolar concentrations) for propylthiouracil were: 7.1 for rT₃ 5'MDL, 1.5 for rT₃ 5'MDH, 24 for T₄ 5'MDL, and 40 for T₄ 5'MDH; those for iopanoate were 0.4 for rT₃ 5'MDL, 18 for rT₃ 5'MDH, 7.0 for T₄ 5'MDL, and 4.0 for T₄ 5'MDH. DTT was a potent stimulator of enzyme activities studied; its dose (minimolar concentrations) that caused a 50% maximal stimulation was 0.04 for rT₃ 5'MDL, 1.0 for rT₃ 5'MDH, 0.025 for T₄ 5'MDL, and 0.035 for T₄ 5'MDH. T₄ inhibited rT₃ 5'-monodeiodination and vice versa. The K_i of T₄ was 1.3 MM for rT₃ 5'MDL and 2.0 for rT₃ 5'MDH, while that of rT₃ was 0.4 for T₄ 5'MDL and 0.6 for T₄ 5'DH. We examined the activity of the hepatic 5'MDL (rT₃, 0.5 nM; DTT, 0.06 nM; 2 M NaCl) and 5'MDH (rT_3, 0.5 µM; DTT, 20 mM; no NaCl) in groups (six animals per group) of rats that were saline treated (control), thyroidectomized, or hyperthyroid (given T_3,20 µg/day for 5 days). The relative values for 5'MDL were (mean ± SD) 17 ± 3.0, 4.0 ± 2.0 (P < 0.01), and 24 ± 2.0 (P < 0.01) pmol/h-mg protein, respectively, whereas those for 5'MDH were 13 ± 4.0, 3.0 ± 1.6 (P < 0.01), and 25 ± 1.0 (P< 0.01) nmol/h · mg protein, respectively.

We conclude that 1) liver contains a low K_m iodothyronine 5'MD which may contribute importantly to T₃ production (from T₄) and rT₃ degradation at physiological substrate concentrations. 2) Activities of hepatic 5'MDL and 5'MDH are similarly inhibited by PTU and affected by thyroid dysfunction; these features of 5'MDL are distinct from those of the 5'MDL described previously in brown fat, cerebral cortex, or anterior pituitary. (Endocrinology 124: 2245-2251, 1989)

Footnotes

^* This work was supported by USPHS Grants AM-16155 from the NIH (Bethesda, MD) and Fogarty International Fellowship 1F05-TW- 03618-01 (to. R.J.B.).

To whom requests for reprints should be addressed

Received November 23, 1988.