Biochemical insights into a novel thermo/organo tolerant bilirubin oxidase from Thermosediminibacter oceani and its application in dye decolorization

Abstract

Abstract Higher activity and stability at neutral pH and tolerance toward anions have made bilirubin oxidases (BODs) proper candidates for industrial utilizations. A putative BOD from Thermosediminibacter oceani (ToBOD) exhibited high stability over a pH range of 3.5–10.0 after heterologous expression and purification. The optimal temperature for the enzyme activity was 75 °C. ToBOD displayed a high thermostability with a half-life of 180 min at 70 °C and 120 min at 80 °C, respectively. Km and Kcat values were 126.5 μM and 130.9 S−1 for ABTS, 19.6 μM and 72.5 S−1 for SGZ, and 31.2 μM and 76.2 S−1 for unconjugated bilirubin, respectively. ToBOD showed tolerance to 10% and 50% (v/v) of water-miscible organic solvents and Triton X-100 as a non-ionic surfactant. In the presence of ABTS as the mediator, ToBOD decolorized malachite green (MG) and Congo red (CR) dyes at a rate of about 63% and 71%, respectively in 2 h. Decolorization was improved within 4 h at a rate of 86% and 89% for MG and CR, respectively. Structural analyses of ToBOD showed that lower folding heat capacity, folding enthalpy, folding free energy, and side-chain hydrogen bonds have a correlation with in vitro biochemical properties.