Purification and characterization glutathione S-transferase enzyme from quail (Coturnix, coturnix japonica) heart and investigation the effect of some metal ions on enzyme activity

Abstract

In this study glutathione S-transferase enzyme (EC: 2.5.1.18) from the heart of japonica quail was purified with 34.0 EU/mg specific activity, 10.44% purification yield and 78.29 purification folds and characterized. Purification processes are consist of three steps, firstly homogenate was prepared, and then ammonium sulfate precipitation was performed and finally glutathione-agarose gel affinity column chromatography was performed. To check the purity of GST enzyme used SDS-PAGE method. Then the M.W calculated at approximately 26.3 kDa by SDS-PAGE method. Enzymatic activity was determined spectrofotometrically according to Beutler`s method at 340 nm. Also characterizations study carry out, and the results obtained are stability-pH = 9.0 in Tris/HCL buffer, optimum pH = 8.0 in Tris/HCl buffer, optimum temperature 60 °C, optimum ionic strength was 1.2 M in Tris/HCl buffer. And kinetic studies performed for GST enzyme purified from quail heart by used both glutathione and 1-chloro 2,4-dinitrobenzen as substrate. KM and Vmax values are determined as 1.642 mM and 0.502 EU/mL respectively for GSH substrate and 3.880 mM and 0.588 EU/mL respectively for CDNB substrate. In addition, the effect of some metal ions (Cu 2+ , Cd 2+ , Fe 2+ , Fe 3+ Zn 2+ , Ag + , Co 2+ , and Ti 1+) were investigated on the GST enzyme activity in vitro.

Bazı Antibiyotiklerin Koyun Dalak Glutatyon S-Transferaz Enzimi Üzerine İn Vitro Etkilerinin İncelenmesi

“…Örneğin Balcı ve arkadaşları bazı pestisitlerin yaban mersini GST enzimi üzerine etkilerini incelenmiş ve bu pestisitlerin enzimi önemli derecede inhibe ettiklerini tespit etmişlerdir [20]. Yine Ahmed ve arkadaşları da bıldırcın kalbinden saflaştırılan GST enzimi üzerine bazı metal iyonlarının etkilerini incelemiş ve bu metal iyonlarından Cd 2+ , Cu 2+ ve Ag + iyonlarının enzimi inhibe ettiklerini bulmuşlardır [23]. Temel ve arkadaşları da bıldırcın karaciğerinden saflaştırılan GST üzerine bazı önemli organik bileşiklerin etkilerini araştırmış ve bu organik bileşiklerin GST enzimini inhibe ettiklerini ortaya koymuşlardır [4].…”

Acute toxicity and metabolism of pesticides in birds

“…95) Similar enzymology, but with homodimers, were reported for the hepatic GSTs of the Japanese quail. 96,97) Incidentally, AFB 1 is more toxic to birds than mammals due to their lack of the ability to metabolize the toxic metabolite AFBO by GST. 62,63) The recombinant α-and μ-GSTs prepared by cloning the corresponding genes of the turkey well reproduced their activity against CDNB and DCNB.…”

Partial Purification of Glutathione S-transferase Enzyme From the Seed of Mallow (Malva Slyvestris L.) and Investigation of the Inhibition Kinetics of Some Heavy Metals

“…The IC 50 values of these metals were 0.0028, 0.382 and 3.127 mM, respectively. The strongest inhibitory metal was found to be Cu 2+[34]. In the study of Akkemik et al 2012, GST enzyme was puri ed from turkey liver and heavy metal inhibition was investigated.…”