title: A role for glutathione transferase Omega 1 (GSTO1-1) in the glutathionylation cycle.
abstract: The glutathionylation of intracellular protein thiols can protect against irreversible oxidation and can act as a redox switch regulating metabolic pathways. In this study we discovered that the Omega class glutathione transferase GSTO1-1 plays a significant role in the glutathionylation cycle. The catalytic activity of GSTO1-1 was determined in vitro by assaying the deglutathionylation of a synthetic peptide by tryptophan fluorescence quenching and in T47-D epithelial breast cancer cells by both immunoblotting and the direct determination of total glutathionylation. Mutating the active site cysteine residue (Cys-32) ablated the deglutathionylating activity of GSTO1-1. Furthermore, we demonstrate that the expression of GSTO1-1 in T47-D cells that are devoid of endogenous GSTO1-1 resulted in a 50% reduction in total glutathionylation levels. Mass spectrometry and immunoprecipitation identified β-actin as a protein that is specifically deglutathionylated by GSTO1-1 in T47-D cells. In contrast to the deglutathionylation activity, we also found that GSTO1-1 is associated with the rapid glutathionylation of cellular proteins when the cells are exposed to S-nitrosoglutathione. The common A140D genetic polymorphism in GSTO1 was found to have significant effects on the kinetics of both the deglutathionylation and glutathionylation reactions. Genetic variation in GSTO1-1 has been associated with a range of diseases, and the discovery that a frequent GSTO1-1 polymorphism affects glutathionylation cycle reactions reveals a common mechanism where it can act on multiple proteins and pathways.
authors.affiliation: From the Department of Molecular Bioscience, John Curtin School of Medical Research, Australian National University, Canberra ACT-2600, Australia.
authors.forename: Deepthi
authors.initials: D
authors.lastname: Menon
authors.affiliation: From the Department of Molecular Bioscience, John Curtin School of Medical Research, Australian National University, Canberra ACT-2600, Australia. Electronic address: Philip.Board@anu.edu.au.
authors.forename: Philip G
authors.initials: PG
authors.lastname: Board
mesh_terms: D000199:Actins / Q000235:genetics / Q000378:metabolism; D019943:Amino Acid Substitution; D020134:Catalytic Domain; D045744:Cell Line, Tumor; D005978:Glutathione / Q000235:genetics / Q000378:metabolism*; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D020125:Mutation, Missense; D011110:Polymorphism, Genetic; D011499:Protein Processing, Post-Translational / Q000502:physiology*
chemical_list: D000199:Actins; C490476:GSTO1 protein, human; D005982:Glutathione Transferase; D005978:Glutathione
keywords: Enzyme Catalysis; Genetic Polymorphism; Glutathione Transferase Omega; Glutathionylation; Redox Regulation; Thiol