Discovery and Characterization of a Second Mammalian Thiol Dioxygenase, Cysteamine Dioxygenase

Abstract

There are only two known thiol dioxygenase activities in mammals, and they are ascribed to the enzymes cysteine dioxygenase (CDO) and cysteamine (2-aminoethanethiol) dioxygenase (ADO). Although many studies have been dedicated to CDO, resulting in the identification of its gene and even characterization of the tertiary structure of the protein, relatively little is known about cysteamine dioxygenase. The failure to identify the gene for this protein has significantly hampered our understanding of the metabolism of cysteamine, a product of the constitutive degradation of coenzyme A, and the synthesis of taurine, the final product of cysteamine oxidation and the second most abundant amino acid in mammalian tissues. In this study we identified a hypothetical murine protein homolog of CDO (hereafter called ADO) that is encoded by the gene Gm237 and belongs to the DUF1637 protein family. When expressed as a recombinant protein, ADO exhibited significant cysteamine dioxygenase activity in vitro. The reaction was highly specific for cysteamine; cysteine was not oxidized by the enzyme, and structurally related compounds were not competitive inhibitors of the reaction. When overexpressed in HepG2/C3A cells, ADO increased the production of hypotaurine from cysteamine. Similarly, when endogenous expression of the human ADO ortholog C10orf22 in HepG2/C3A cells was reduced by RNAmediated interference, hypotaurine production decreased. Western blots of murine tissues with an antibody developed against ADO showed that the protein is ubiquitously expressed with the highest levels in brain, heart, and skeletal muscle. Overall, these data suggest that ADO is responsible for endogenous cysteamine dioxygenase activity.There are many different processes in mammalian cells that result in the oxidation of thiol groups. Because of their reactivity, free sulfhydryl groups are highly susceptible to oxidation that results in the formation of disulfides, sulfenates, sulfinates, and sulfonates. Many of these reactions occur nonenzymatically, principally as a consequence of adventitious free radicals arising from aerobic respiration. Nevertheless, there are a small number of thiol oxidation reactions that are known to occur directly via enzymatic catalysis. The enzymes that catalyze these reactions show a high degree of substrate specificity and confer to cells the advantage of being able to precisely regulate the level of a particular reduced thiol.One interesting subset of the enzymes capable of specifically oxidizing free sulfhydryl groups are the thiol dioxygenases. In mammals this family comprises only two known proteins: cysteine dioxygenase (CDO, 3 EC 1.13.11.20) and cysteamine dioxygenase (EC 1.13.11.19). CDO adds two atoms of oxygen to free cysteine to yield cysteine sulfinic acid, whereas cysteamine dioxygenase adds two atoms of oxygen to free cysteamine (2-aminoethanethiol) to form hypotaurine (Fig. 1). The activities for these two proteins were first reported in mammalian tissues almost 40 years ago (1, 2). Since that time, howev...

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