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The synthesis of adrenodoxin, a mitochondrial iron-sulfur protein required for adrenocortical steroidogenesis, is known to be regulated chronically by ACTH. Rhodanese, also a mitochondrial enzyme, is thought to be required for synthesis of iron-sulfur centers, such as those contained in adrenodoxin. In this study it has been found that rhodanese synthesis and activity are not regulated by ACTH, under the same conditions whereby ACTH induces adrenodoxin synthesis. In addition, unlike adrenodoxin, rhodanese is found to be synthesized in the mature form rather than as a higher molecular weight precursor protein.