Interactions of Sodium Selenite, Glutathione, Arsenic Species, and Omega Class Human Glutathione Transferase

• Published in: Chemical research in toxicology Vol. 18; no. 8; pp. 1287 - 1295
• Main Authors: Zakharyan, Robert A, Tsaprailis, George, Chowdhury, Uttam K, Hernandez, Alba, Aposhian, H. Vasken
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 01-08-2005
• Subjects: Amino Acid Sequence; Arsenicals - pharmacology; Cysteine - chemistry; Dithiothreitol - pharmacology; Enzyme Inhibitors - pharmacology; Glutathione - pharmacology; Glutathione Transferase - antagonists & inhibitors; Glutathione Transferase - metabolism; Humans; Kinetics; Molecular Sequence Data; Oxidation-Reduction; Reactive Oxygen Species; Recombinant Proteins - chemistry; Sodium Selenite - antagonists & inhibitors; Sodium Selenite - pharmacology; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Stereoisomerism; Sulfhydryl Compounds - chemistry; Sulfhydryl Compounds - metabolism
• ISSN: 0893-228X; 1520-5010
• DOI: 10.1021/tx0500530

Human monomethylarsenate reductase [MMA(V) reductase] and human glutathione S-transferase omega 1-1 (hGSTO1-1) [because MMA(V) reductase and hGSTO1-1 are identical proteins, the authors will utilize the designation “hGSTO1-1”] are identical proteins that catalyze the reduction of arsenate, monomethylarsenate [MMA(V)], and dimethylarsenate [DMA(V)]. Sodium selenite (selenite) inhibited the reduction of each of these substrates by the enzyme in a concentration-dependent manner. The kinetics indicated a noncompetitive inhibition of the MMA(V), DMA(V), or arsenate reducing activity of hGSTO1-1. The inhibition of the MMA(V) reducting activity of hGSTO1-1 by selenite was reversed by 1 mM dl-dithiothreitol (DTT) but not by reduced glutathione (GSH), which is a required substrate for the enzyme. Neither superoxide anion nor hydrogen peroxide was involved in the selenite inhibition of hGSTO1-1. MALDI-TOF and MS/MS analysis demonstrated that five molecules of GSH were bound to one monomer of hGSTO1-1. Four of the five cysteines of the monomer were glutathionylated. Cys-32 in the active center, however, exists mostly in the sulfhydryl form since it was alkylated consistently by iodoacetamide. MALDI-TOF mass spectra analysis of hGSTO1-1 after reaction with GSH and sodium selenite indicated that selenium was integrated into hGSTO1-1 molecules. Three selenium were found to be covalently bonded to the monomer of hGSTO1-1 with three molecules of GSH. It is proposed that the reaction products of the reduction of selenite inhibited the activity of hGSTO1-1 by reacting with disulfides of glutathionylated cysteines to form bis (S-cysteinyl)selenide and S-selanylcysteine and had little or no interaction with the sulfhydryl of Cys-32 in the active site of the enzyme.