Investigation of the effects of purification and characterization of turkey (Meleagris gallopavo) liver mitochondrial thioredoxin reductase enzyme and some metal ions on enzyme activity


Abstract: 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 $\times $ mg$^{-1}$ specific activity was purified 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 filtration chromatography. In addition, characteristic and kinetic properties of the enzyme were determined. Then the inhibitory effects of some heavy metal ions (Ag$^{+}$, Fe$^{3+}$, Cd$^{2+}$, Cu$^{2+}$, Zn$^{2+}$, Pb$^{2+}$, Ni$^{2+}$, and Co$^{2+})$ on the activity of TrxR enzyme were examined under in vitro conditions. IC$_{50}$ values were found with the heavy metal concentration with which 50{\%} of the activity of the TrxR enzyme was inhibited. Finally, K$_{i}$ values for these substances were calculated from the Lineweaver--Burk plots. It was determined that Ag$^{+}$, Fe$^{3+}$, Cd$^{2+}$, Cu$^{2+}$, and Zn$^{2+}$ ions inhibited TrxR enzyme, Pb$^{2+}$ ion increased enzyme activity, and Ni$^{2+}$ and Co$^{2+}$ ions had no effect on enzyme activity.

Keywords: Thioredoxin reductase, characterization, metal, inhibition

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