Purification and characterization of metmyoglobin reductase from ordinary muscle of blue-fin tuna

Abstract

Metmyoglobin (Met-Mb) reductase from blue-fin tuna ordinary muscle was purified to electrophoretical homogeneity by ammonium sulfate fractionation, ion exchange and organomercurial agarose affinity chromatography. The molecular weight estimated by SDS-PAGE was 100 kDa. The optimal pH and temperature for the reduction of met-Mb were 7.3 and 25°C, respectively. This enzyme was very stable at pH 7.0-7.3 and 4°C-15°C. The purified enzyme was strongly activated by K+, moderately activated by Na+ and Mn2+, but not affected by Ni2+. It was moderately inhibited by Li+, NH4+, Mg2+, and Co2+. The Vmax for the reduction of met-Mb and NADH were 0.32 and 0.43 mM/min/mg, while Km were 2.3 × 10-5 and 24.4 × 10-5 M, respectively.