Molecular delineation, immune modulation analysis, and antioxidant functional assessment of catalase (CAT) from chub...

Cytotoxic reactive oxygen species (ROS), including hydroxyl radicals (^.OH^-), superoxide anions (^.O₂^-), peroxides (^.O₂^2-), and nitrogen radicals (NO), are derivatives of hydrogen peroxide (H₂O₂). Catalase (CAT) is an immunologically important antioxidant enzyme that regulates host redox homeostasis by disintegrating cellular H₂O₂ into water and oxygen. The identified CAT homolog sequence from Scomber japonicus (SjCAT) comprised 1584 bp, encoding 527 amino acids in length with a molecular weight of 59.99 kDa, and an isoelectric point of 7.69. In silico analysis revealed that the SjCAT sequence exhibited the highest evolutionary identity (96 %) and similarity (97.9 %) with the Thunnus maccoyii ortholog. The CAT orthologs possessed a highly conserved characteristic active site signature motif, and a heme-binding ligand, consistent with findings among teleost fish and other identified vertebrate counterparts. The highest tissue-specific SjCAT mRNA expression was observed in blood, followed by the brain, heart, muscle, and liver. Furthermore, significant modulation of SjCAT expression was observed in the blood and spleen after treatment with polyinosinic: polycytidylic acids, lipopolysaccharide, Vibrio harveyi, and Streptococcus iniae immunostimulants. The antioxidant activity of the recombinant SjCAT protein was determined by analyzing its activity under varying pH and temperature, and its H₂O₂ radical scavenging activity. Furthermore, cell-based assays demonstrated that SjCAT provides multiple protective effects, including enhancing cell viability, suppressing apoptosis under H₂O₂ stress, scavenging NO, and reducing metal cation-induced ROS production. These findings confirmed its antioxidant function and highlight its potential to support immune cell survival during oxidative stress associated with immune responses. Overall, these results suggest that SjCAT plays vital antioxidant and immunological roles, contributing to the survival of chub mackerel in redox-unstable environments and during pathogenic challenges.