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Glutathione-S-transferase (GSTs) is a multifunctional enzyme that provides homeostasis by catalyzing the first step of the formation of mercapturic acid, the end product in detoxification metabolism. They can prevent reactive electrophilic compounds from harming the body by covalently binding the same type of compounds to each other. In this study, we determined the in vitro inhibitory effects of metal ions such as Cu2+, Cd2+, Ag+, and Co2+ on GST enzyme activity. For this aim, GST was purified from C. tarichi Pallas liver with 37.36% yield and 29.304 EU/mg specific activity using the chromatographic method. The Vmax values of liver GST were determined for CDNB and GSH to be 1.245 and 0.562 EU/mL, respectively, and the Km values were found 0.89 and 0.06 mM, respectively, using the Lineweaver-Burk plot. The effects of the metal ions at different concentrations on in vitro GST activity were studied. The IC50 values were determined for Cu+2, Cd+2, Ag+, and Co+2 as 0.163, 0.235, 0.00021, and 0.446 mM, respectively. The Ki constants were determined as 0.049 ± 0.009, 0.117 ± 0.031, 0.002 ± 0.0007, and 0.893 ± 0.3 mM, respectively. Ag+ showed the best inhibitory effect among the studied metal ions. Cd2+, Cu2+ and Co2+ showed a competitive inhibition mechanism, while Ag+ was noncompetitive. [ABSTRACT FROM AUTHOR]

NADPH-Cytochrome P450 reductase (CPR) is a key enzyme catalyzing electron transfer in detoxification metabolism. In this study, two methods were employed to purify CPR enzyme from turkey liver microsomes. In the first method, NADPH-Cytochrome P450 reductase was purified using 2ʹ, 5ʹ-ADP Sepharose 4B affinity column with ~114 purification fold and ~23% yield. In the second method, CPR was purified using DE-52 Cellulose anion exchange column and 2ʹ, 5ʹ-ADP Sepharose 4B affinity column with 124 purification fold and 8% yield. Enzyme purity was checked in both methods with SDS-PAGE. Characteristics kinetic features of the purified enzyme were identified. The effects of some metal ions on purified PCR enzyme activity have been investigated in vitro conditions. It has been found that Ag+, Hg2+ and Cu2+ metal ions have an inhibitory effect on CPR enzyme activity. [ABSTRACT FROM AUTHOR]

The thioredoxin system, found in all living creatures, consists of thioredoxin protein (Trx), thioredoxin reductase enzyme (TrxR), and NADPH. In this study, turkey liver mitochondrial TrxR enzyme with 3.07 EU × mg-1 speci c activity was puri ed 990-fold in a yield of 2.05% using 2',5'-ADP Sepharose 4B affinity chromatography. The purity of the enzyme was measured and the molecular weight of its subunits was determined to be 45.5 kDa by SDS- PAGE. The molecular mass of the enzyme's natural state was found to be 88 kDa using Sephadex-G 150 gel ltration chromatography. In addition, characteristic and kinetic properties of the enzyme were determined. Then the inhibitory effects of some heavy metal ions (Ag+, Fe3+, Cd2+, Cu2+, Zn2+, Pb2+, Ni2+, and Co2+) on the activity of TrxR enzyme were examined under in vitro conditions. IC50 values were found with the heavy metal concentration with which 50% of the activity of the TrxR enzyme was inhibited. Finally, Ki values for these substances were calculated from the Lineweaver-Burk plots. It was determined that Ag+, Fe3+, Cd2+, Cu2+, and Zn2+ ions inhibited TrxR enzyme, Pb2+ ion increased enzyme activity, and Ni2+ and Co2+ ions had no effect on enzyme activity. [ABSTRACT FROM AUTHOR]

Thioredoxin reductase (E.C 1.6.4.5.; TrxR) is an enzyme belonging to the flavoprotein family of pyridine nucleotide-disulfide oxidoreductases. In this study, mitochondrial TrxR enzyme was purified from rainbow trout mitochondria. Thanks to the 2 consecutive procedures (preparation of homogenate and 2',5'-ADP Sepharose 4B affinity chromatography), the enzyme, having the specific activity of 11.9 EU mg protein-1, was purified with a yield of 2.38% and 672-fold. The purity of the enzyme was monitored and the molecular weight of its subunits was calculated as 70 kDa by SDS-PAGE. The native molecular mass of the enzyme was found to be approximately 151 kDa by gel filtration chromatography. Characteristic and kinetic properties of the enzyme were also determined. Furthermore, Se4+ , Cu2+ , Co2+ , Ni2+ , Fe3+ , and Al3+ metal ions' in vitro effects on mitochondrial TrxR purified from rainbow trout was investi- gated. While Se4+ ion increased the enzyme activity, all of the other metal ions used in this study showed an inhibitory effect. [ABSTRACT FROM AUTHOR]