Purification and partial characterization of thioredoxin reductase from the hepatopancreas of the mollusc Mytilus galloprovincialis Lam.

Yıl 2018, Cilt: 48 Sayı: 1, 12 - 17, 30.04.2018

Esra Acar Gözde Hasbal Nurten Özsoy

Öz

DOI: 10.5152/IstanbulJPharm.2018.382627

Thioredoxin reductase (TrxR, EC 1.6.4.5) is
a ubiquitous flavoenzyme that is present from Archaea to humans, and it is the
only enzyme capable of catalyzing the reduction of thioredoxin (Trx) by nicotinamide
adenine dinucleotide phosphate (NADPH). Although TrxR has been purified and
characterized from different bacteria, plants, and mammalian organisms, a
survey of the literature revealed no studies on the purification and
characterization of TrxR from the mussel Mytilus galloprovincialis Lam. In this
study, TrxR was purified to homogeneity from the hepatopancreatic tissue of M.
galloprovincialis Lam. by extraction, ammonium sulfate precipitation, and
DEAE-Sepharose CL-6B anion and 2’,5‘-ADP-agarose chromatographies, and some of
its kinetic properties were examined. Molar mass determined by sodium dodecyl
sulfate polyacrylamide gel electrophoresis revealed only a single protein band
corresponding to a molecular weight of 35 kDa. Optimum pH and temperature were
found to be 7.0 and 60°C, respectively. Km and Vmax values for NADPH were found
to be 85 µmol and 4.82 µmol/min/mg, respectively. For 5,5ʹ-dithiobis
(2-nitrobenzoic) acid (DTNB), the Km and Vmax values were 193 µmol and 1.32
µmol/min/mg, respectively. Increasing the knowledge on the kinetic properties
of TrxR will significantly increase the prospects of enzyme application as an
oxidative stress biomarker in mussels and fishes for monitoring contamination
in coastal environments.

Anahtar Kelimeler

Enzyme purification, kinetic properties, Mytilus galloprovincialis Lam.

Kaynakça

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