title: Electrochemical evaluation of glutathione S-transferase kinetic parameters.
abstract: Glutathione S-transferases (GSTs), are a family of enzymes belonging to the phase II metabolism that catalyse the formation of thioether conjugates between the endogenous tripeptide glutathione and xenobiotic compounds. The voltammetric behaviour of glutathione (GSH), 1-chloro-2,4-dinitrobenzene (CDNB) and glutathione S-transferase (GST), as well as the catalytic conjugation reaction of GSH to CDNB by GST was investigated at room temperature, T=298.15K (25°C), at pH6.5, for low concentration of substrates and enzyme, using differential pulse (DP) voltammetry at a glassy carbon electrode. Only GSH can be oxidized; a sensitivity of 0.14nA/μM and a LOD of 6.4μM were obtained. The GST kinetic parameter electrochemical evaluation, in relation to its substrates, GSH and CDNB, using reciprocal Michaelis-Menten and Lineweaver-Burk double reciprocal plots, was determined. A value of KM~100μM was obtained for either GSH or CDNB, and Vmax varied between 40 and 60μmol/min per mg of GST.
authors.affiliation: Department of Chemistry, University of Coimbra, 3004-535 Coimbra, Portugal.
authors.forename: Teodor Adrian
authors.initials: TA
authors.lastname: Enache
authors.affiliation: Department of Chemistry, University of Coimbra, 3004-535 Coimbra, Portugal. Electronic address: brett@ci.uc.pt.
authors.forename: Ana Maria
authors.initials: AM
authors.lastname: Oliveira-Brett
mesh_terms: D004137:Dinitrochlorobenzene / Q000737:chemistry / Q000378:metabolism; D055664:Electrochemical Techniques; D005978:Glutathione / Q000737:chemistry / Q000378:metabolism; D005982:Glutathione Transferase / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D010084:Oxidation-Reduction
chemical_list: D004137:Dinitrochlorobenzene; D005982:Glutathione Transferase; D005978:Glutathione
keywords: 1-Chloro-2,4-dinitrobenzene; Electrochemistry; Glutathione; Glutathione S-transferase; Oxidation