Journal of Agriculture and Crops ISSN(e): 2412-6381, ISSN(p): 2413-886X Vol. 2, No. 10, pp: 94-100, 2016 URL: http://arpgweb.com/?ic=journal&journal=14&info=aims *Corresponding Author 94 Academic Research Publishing Group Some Kinetic Properties and Inhibition of Glutathione S- Transferase from a Hybridized Wheat (Triticum aestivum L.) Hülya Öztürk Doğan Atatürk University, Erzurum Vocational College, Chemistry and Chemical Processing Technologies, 25240 Erzurum-Turkey Mustafa Erat * Atatürk University, Erzurum Vocational College, Chemistry and Chemical Processing Technologies, 25240 Erzurum-Turkey 1. Introduction Glutathione S-transferases (GSTs, EC 2.5.1.18) are a family of multifunctional enzymes involved in the detoxification processes through several different mechanisms. These proteins detoxify electrophilic xenobiotics and endogenous compounds by catalysing their conjugation with the tripeptide glutathione to the electrophilic centre of lipophilic compounds, thereby increasing their solubility and removing toxic compounds from circulation through covalent and non-covalent binding [1-3]. The enzymes are also involved in several additional functions including the binding, transport, storage of hydrophobic ligands, [4] the isomerization of maleylacetoacetate [5] and the regulation of stress kinases and apoptosis.[6] Furthermore, these proteins are known to protect cellular integrity from endogenous oxidative stress by expressing selenium–independent glutathione peroxidase activity with organic hydroperoxides. [6] Plant GSTs are a multigene family of enzymes that constitute about 1% of the soluble protein in photosynthetic plant cells. [7] These enzymes are known to function in herbicide detoxification, tyrosine metabolism, hormone homeostasis, vacuolar sequestration of anthocyanin, hydroxyperoxide detoxification, regulation of apoptosis and in plant responses to biotic and abiotic stresses [8]. Glutathione S-transferase and its izozymes have been extensively purified from different sources such as plant, [9, 10] mammalian [11] and microbial sources [12, 13]. In order to purify the enzymes, various chromatographic techniques such as, affinity, ion-exchange, reversed phase, hydrophobic, and size-exclusion columns were used in different reports [14, 15]. In the current study, we partially purified glutathione S-transferase from a hybridized wheat leaves and searched for its some kinetic and characteristic properties. We identified its different properties from the other GST enzymes purified and characterized so far from the other sources. 2. Experimental 2.1. Materials Wheat (Triticum aestivum L.), used in this study was harvested from a field in Erzurum, Turkey. Sephadex G- 100 used as gel filtration column, substrates, and other chemicals were purchased from Sigma Chem. Co. All chemicals used in this study were the best grade available. Substrates and other solutions were prepared freshly in distilled water. 2.2. Enzyme Extraction and Purification 15 grams of wheat samples were cleaned and prepared for the extraction. The samples were immersed in liquid nitrogen, in a Dewar flask to disrupt cell membranes, and were homogenized in 75 ml of 0.1 M potassium phosphate buffer pH 6.0. The crude extract samples were centrifuged at 14,000g for 30 min. The process was conducted at 4 o C. The supernatant was precipitated at 0-20%, 20-40%, 40-60% and 60-80% neutral salt concentrations to find Abstract: Glutathione S-transferase enzymes (GSTs) play central roles in phase II detoxification of both xenobiotics and endogenous compounds in almost all living organisms. The enzyme was extracted and partially purified from wheat leaves through a procedure including ammonium sulfate fractionation followed by dialysis and gel filtration chromatography. These procedures yielded a 7.14-fold purification with 71% recovery. Optimum activity conditions-pH, temperature and ionic strength-of the enzyme were determined. Its some kinetic properties such as Vmax, KM, and kcat were calculated for GSH and CDNB substrates. The kcat/KM values of the enzyme were 603.5 for GSH and 385.3 for CDNB. The native molecular weight of the enzyme was estimated to be 52 kDa based on its mobility in gel filtration column. Keywords: Wheat; Glutathione S-transferase; Purification; Kinetic.