Purification and characterization of mitochondrial thioredoxin reductase enzyme from rainbow trout (Oncorhynchus mykiss) liver and investigation of the in vitro effects of some metal ions on the enzyme


Abstract: Thioredoxin reductase (E.C; 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, Se$^{4+}$, Cu$^{2+}$, Co$^{2+}$, Ni$^{2+}$, Fe$^{3+}$, and Al$^{3+}$ metal ions' in vitro effects on mitochondrial TrxR purified from rainbow trout was investigated. While Se$^{4+}$ ion increased the enzyme activity, all of the other metal ions used in this study showed an inhibitory effect.

Keywords: Thioredoxin reductase, characterization, purification, rainbow trout, metal ions

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