In vitro susceptibility of grass carp fish (Ctenopharyngodon idella) to oxidized myosin on structural change of protein secondary structure associated with reactive oxygen species (ROS) treatment

Abstract

This study aimed to investigate the in vitro susceptibility of grass carp (Ctenopharyngodon idella) oxidized myosin, with the treatment of H2O2 levels being 0, 25, 50, and 100 μmol/L, respectively. The results showed that in vitro oxidation of myosin, reactive oxidative species promoted protein modifications that propagated the conformation changes on myosin secondary structure, leading to a significant increase (P < 0.05) in the protein carbonyl content in the high oxidation treatment, when compared to the low and medium treatments. The results of the Western blot analysis and sodium-polyacrylamide amide-dodecyl sulfate gel electrophoresis demonstrated that in vitro oxidation actively promoted the degradation of protein molecular weight and Desmin (53 kDa) deterioration in the sample with the highest concentration treatment of H2O2. Circular dichroism spectra showed a decrease in α-helix content from 70.1% to 29.8% and an increase in β-sheet percentage from 5.6% to 17.4%. Meanwhile, cysteine and methionine as the free amino acids were most susceptible to oxidative change due to their high sensitivity to the reaction of the sulfur group of amino acids. Protein degradation induced by oxidation resulted in significant changes to the protein secondary structure, as evidenced by the observation of oxidized myosin under different H2O2 levels.