title: Steady-state substrate specificity and O₂-coupling efficiency of mouse cysteine dioxygenase.
abstract: Cysteine dioxygenase (CDO) is a non-heme mononuclear iron enzyme that catalyzes the oxygen-dependent oxidation of L-cysteine (Cys) to produce L-cysteine sulfinic acid (CSA). Sequence alignment of mammalian CDO with recently discovered thiol dioxygenase enzymes suggests that the mononuclear iron site within all enzymes in this class share a common 3-His first coordination sphere. This implies a similar mechanistic paradigm among thiol dioxygenase enzymes. Although steady-state studies were first reported for mammalian CDO over 45 years ago, detailed analysis of the specificity for alternative thiol-bearing substrates and their oxidative coupling efficiencies have not been reported for this enzyme. Assuming a similar mechanistic theme among this class of enzymes, characterization of the CDO substrate specificity may provide valuable insight into substrate-active site intermolecular during thiol oxidation. In this work, the substrate-specificity for wild-type Mus musculus CDO was investigated using NMR spectroscopy and LC-MS for a variety of thiol-bearing substrates. Tandem mass spectrometry was used to confirm dioxygenase activity for each non-native substrate investigated. Steady-state Michaelis-Menten parameters for sulfinic acid product formation and O₂-consumption were compared to establish the coupling efficiency for each reaction. In light of these results, the minimal substrate requirements for CDO catalysis and O₂-activation are discussed.
authors.affiliation: Department of Chemistry and Biochemistry, College of Sciences, The University of Texas at Arlington, Arlington, TX 76019, United States.
authors.forename: Wei
authors.initials: W
authors.lastname: Li
authors.affiliation: Department of Chemistry and Biochemistry, College of Sciences, The University of Texas at Arlington, Arlington, TX 76019, United States. Electronic address: bspierce@uta.edu.
authors.forename: Brad S
authors.initials: BS
authors.lastname: Pierce
mesh_terms: D000818:Animals; D002384:Catalysis; D050558:Cysteine Dioxygenase / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D010084:Oxidation-Reduction; D010100:Oxygen / Q000737:chemistry* / Q000378:metabolism; D013379:Substrate Specificity / Q000502:physiology; D013438:Sulfhydryl Compounds / Q000737:chemistry* / Q000378:metabolism
chemical_list: D013438:Sulfhydryl Compounds; D050558:Cysteine Dioxygenase; D010100:Oxygen
keywords: Coupling; Cysteine; Dioxygenase; Non-heme iron; Specificity; Thiols