A novel highly thermostable and stress resistant ROS scavenging metalloprotein from Paenibacillus

Reactive oxygen species (ROS) are unstable metabolites produced during cellular respiration that can cause extensive damage to the body. Here we report a unique structural metalloprotein called RSAP_p for the first time, which exhibits robust ROS-scavenging activity, high thermostability, and stress resistance. RSAP_p is a previously uncharacterized DUF2935 (domain of unknown function, accession number: cl12705) family protein from Paenibacillus, containing a highly conserved four-helix bundle with binding sites for variable-valence metal ions (Mn²⁺/Fe²⁺/Zn²⁺). Enzymatic characterization results indicated that RSAP_p displays the functionality of three different antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD). In particular, RSAP_p exhibits a significant SOD-like activity that is remarkably effective in eliminating superoxide radicals (up to k_cat/K_M = 2.27 × 10¹¹ mol^-1 s^-1), and maintains the catalytical active in a wide range of temperatures (25-100 °C) and pH (pH 2.0-9.0), as well as resistant to high temperature, alkali and acidic pH, and 55 different concentrations of detergent agents, chemical solvents, and inhibitors. These properties make RSAP_p an attractive candidate for various industrial applications, including cosmetics, food, and pharmaceuticals.