Structure-specific effects of thyroxine analogs on human liver 3α-hydroxysteroid dehydrogenase

The NADP(H)-linked oxidoreductase activity of a major isozyme of human liver 3 alpha-hydroxysteroid dehydrogenase was activated 5-, 4-, and 2-fold by D-thyroxine (T-4), L-T-4 and DL-3,3',5'-triiodothyronine (reverse T-3), respectively. Kinetic analysis of the activation indicated that D-T-4, L-T-4, and reverse T-3 are non-essential activators, showing binding constants of 1.5, 1.1, and 3.6 mu M, respectively. Comparison of the effects of the T-4 analogs on the activities of the mutant enzymes suggests that the binding site is composed of at least Lys-270, Arg-276, and the C-terminal loop of the enzyme. L-T-3, DL-thyronine, and D-tyrosine had no effect on the enzyme, but 3,5,3',5'-tetra- and 3,5,3'-tri-iodothyropropionic acids were potent competitive inhibitors with K-i values of 42 and 60 nM, respectively, with respect to the substrate. The inhibition constant was lowered upon the activation of the enzyme by D-T-4, and the inhibition by the deamino derivatives of T-4 and T-3 disappeared upon modification of the C-terminal loop of the enzyme, but not upon replacement of Lys-270 or Arg-276 with Met. These results indicate that, depending on their structures, the T-4 analogs bind differently to two distinct sites at the active center of the enzyme to produce stimulatory and inhibitory effects.