Characterization of hepatic low-K(m) outer-ring deiodination in red drum (Sciaenops ocellatus).
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abstract
The more biologically active thyroid hormone 3,5,3'-triiodothyronine (T(3)), is primarily derived from peripheral deiodination of thyroxine (T(4)). We characterized hepatic deiodination for a commercially important, warm water teleost fish, the red drum (Sciaenops ocellatus). Low K(m) outer-ring deiodination (ORD) activity was determined by production of free iodide ((125)I) upon incubation of hepatic microsomes with radiolabeled T(4). HPLC analysis demonstrated that (125)I, and T(3) were produced in equal amounts, thereby validating 125I as a measure of T(3) production. A small amount of 3,3',5'-triiodothyronine (reverse T(3)) was also produced by inner-ring deiodination. Production of (125)I was linear over a range of 0--100 microg protein/ml and for incubations of 30 min--4 h. Maximal ORD activity was measured at pH 6.6, 50 mM dithiothreitol (DTT) and an incubation temperature of 20 degrees C. Double reciprocal plots demonstrated that the average apparent K(m) was 5.1 nM and the average V(max) was 3.7 pmol T(4) converted/h per mg protein. ORD was not inhibited by propylthiouracil but was 50% inhibited by 90 microM of iodoacetic acid and 7 microM of gold thioglucose. The substrate analog preference was T(4) = tetraiodoacetic acid = reverse T(3) > triiodoacetic acid >> T(3). In relation to other tissues, ORD for liver>gill>intestine>kidney. Similar hepatic deiodination activity was present in adult wild, aquacultured and laboratory-reared red drum, but in adult wild red drum the optimum temperature was higher. Red drum hepatic low-K(m) deiodination activity appears to most closely resemble rainbow trout hepatic and mammalian Type II deiodination. Evidence of inner-ring T(4) deiodination suggests a more active hepatic iodothyronine catabolic pathway than in other teleost species.
