A tale of sulfur and selenium metabolites: From dioxygenases with sulfur-containing substrates to the analysis of selenium by gas chromatography with atomic emission detection
Sulfur and selenium participate in a variety of essential biological functions. The investigation of two non-heme iron dioxygenases with sulfur-containing substrates is reported along with studies of seleno-compounds by gas chromatography with atomic emission detection (GC-AED).
Cysteine dioxygenase (CDO) 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 E. coli as a 26.8 kDa fusion protein bearing a poly-histidine tag. Kinetics studies revealed a Km value of 2.5 ± 0.4 mM at pH 7.5 and 37 °C, with no requirement for secondary proteins or cofactors. Fe was demonstrated to be the only metal that is essential for activity. Inhibition studies with cysteine analogs along with the use of x-ray absorption spectroscopy (XAS) allowed for the characterization of the active site.
Acireductone dioxygenases (ARDs) are enzymes involved in the methionine recycle pathway. Klebsiella produces two ARD enzymes that share a common polypeptide sequence and differ only in the metal ion present. In the presence of Fe-containing ARD (ARD'), reaction of acireductone with dioxygen produces formate and the ketoacid precursor of methionine. In the presence of the Ni-bound form (ARD) the same substrate produces formate, methylthiopropionate and CO, an off-pathway shunt. XAS study of the structure of the catalytic Fe center in resting state enzyme shows a six coordinate Fe site composed of N/O-donor ligands including 3-4 histidine residues. The substrate binds to the Fe center in a bidentate fashion by displacing two ligands, at least one of which is a histidine ligand.
Cancer prevention attributed to the properties of selenium is widely recognized, but the mechanism of tumor inhibition by this element is still not known. The determination of Se-metabolites and the understanding of their fate are indispensable. Liver extracts from rats administered with selenized yeast were examined by GC-AED. Organoseleno compounds have been observed, including selenomethionine, and possibly the newly-discovered S-(methylseleno)-cysteine molecule. Aromatic Se-compounds have been recognized as antitumorigenic alternatives with lower toxicity. Gas chromatographic behavior of several aryl diselenide derivatives were analyzed.
0486: Analytical chemistry