Abstract
A novel bilirubin oxidase (BOD), from the rice blast fungus Magnaporthe oryzae, has been identified and isolated. The 64-kDa protein containing four coppers was successfully overexpressed in Pichia pastoris and purified to homogeneity in one step. Protein yield is more than 100 mg for 2 L culture, twice that of Myrothecium verrucaria. The k(cat)/K(m) ratio for conjugated bilirubin (1,513 mM⁻¹ s⁻¹) is higher than that obtained for the BOD from M. verrucaria expressed in native fungus (980 mM⁻¹ s⁻¹), with the lowest K(m) measured for any BOD highly desirable for detection of bilirubin in medical samples. In addition, this protein exhibits a half-life for deactivation >300 min at 37 °C, high stability at pH 7, and high tolerance towards urea, making it an ideal candidate for the elaboration of biofuel cells, powering implantable medical devices. Finally, this new BOD is efficient in decolorizing textile dyes such as Remazol brilliant Blue R, making it useful for environmentally friendly industrial applications.