Cysteine dioxygenase (CDO) catalyzes the oxidation of cysteine to cysteine sulfinic acid, which is the first major step in cysteine catabolism in mammalian tissues. Crystal structures of mouse, rat, human and bacterial CDO have recently become available and provide significant mechanistic insights. Unlike most non-heme Fe(II) dioxygenases, coordination of the Fe in CDO deviates from the 2-His-1-carboxylate facial triad archetype and instead adopts a His₃ facial triad. This change is expected to have an influence on oxygen activation by the catalytic site. The structures also reveal the presence of a cysteinyltyrosine (Tyr157-Cys93) post-translational modification near the active site. Kinetic studies of mutant CDOs reveal that the cysteine residue is less critical than the tyrosine for enzyme activity. Inconsistencies about the details of the active site and the nature of substrate binding exist and are discussed. Herein we review the structural biology along with relevant kinetics studies that have been conducted on CDO for insights into the reaction mechanism of this novel non-heme iron dioxygenase.

Cysteine dioxygenase (CDO, EC 1.13.11.20) catalyzes the oxidation of cysteine to cysteine sulfinic acid, which is the first major step in cysteine catabolism in mammalian tissues. Rat liver CDO was cloned and expressed in Escherichia coli as a 26.8-kDa N-terminal fusion protein bearing a polyhistidine tag. Purification by immobilized metal affinity chromatography yielded homogeneous protein, which was catalytically active even in the absence of the secondary protein-A, which has been reported to be essential for activity in partially purified native preparations. As compared with those existing purification protocols for native CDO, the milder conditions used in the isolation of the recombinant CDO allowed a more controlled study of the properties and activity of CDO, clarifying conflicting findings in the literature. Apo-protein was inactive in catalysis and was only activated by iron. Metal analysis of purified recombinant protein indicated that only 10% of the protein contained iron and that the iron was loosely bound to the protein. Kinetic studies showed that the recombinant enzyme displayed a K_m value of 2.5 ± 0.4 mm at pH 7.5 and 37 °C. The enzyme was shown to be specific for l-cysteine oxidation, whereas homocysteine inhibited CDO activity.