Purification and properties of bilirubin oxidase fromMyrothecium verrucaria

Abstract

Bilirubin oxidase was purified from a culture filtrate ofMyrothecium verrucaria Mv 2,1089 by DEAE-cellulose and Sephadex G-100 column chromatographies. The purified enzyme had a specific activity of 30 U/mg protein and showed a single band on polyacrylamide gel electrophoresis.

Some of the general properties of this bilirubin oxidase were as follows: the optimum pH for the enzyme reaction was 7.5 and the optimum temperature was 50°C. The enzyme was stable at pH ranging from 9.0 to 9.5. The mol wt was calculated to be 61,900–62,700 by SDS-PAGE and gel-filtration technique. The apparentK _m value of the bilirubin oxidase was calculated to be 9.4x10^-5 mol/L. The enzyme activity was greatly reduced by incubation of bilirubin oxidase with Fe²⁺, Hg⁺, NaN₃, NH⁺ ₄ , and Zn²⁺. The enzyme reaction was inhibited in the presence of Ca²⁺, Hg⁺, Zn²⁺, Fe²⁺, and BSA.