Glyoxal oxidase supplies hydrogen peroxide at hyphal tips and on hyphal wall to manganese peroxidase of white-rot fungus Phanerochaete crassa WD1694

Peroxidase activity staining localized at hyphal tips of white-rot fungus Phanerochaete crassa WD1694 that was cultivated in a shaken liquid culture containing unbleached kraft pulp was investigated. Manganese peroxidase was detected in culture solution, washing solution of mycelium, and mycelial extract. Glyoxal oxidase was detected only in mycelial extract and was not detected in culture solution. Addition of hydrogen peroxide generated peroxidase activity staining in the culture solution. Addition of catalase resulted in no staining in the culture of P. crassa WD1694, and the addition of methylglyoxal resulted in marked peroxidase activity staining at hyphal tips and on hyphal wall. In an optimized culture, glyoxal oxidase was produced in culture solution. Although the production of glyoxal oxidase and manganese peroxidase had a positive correlation, the secretion and the peak of glyoxal oxidase was observed 3 and 2 days later than those of manganese peroxidase. The N-terminal sequence of purified glyoxal oxidase had very high homology with that of P. chrysosporium. These results elucidated the hydrogen peroxide supply system in lignin biodegradation by white-rot fungi, i.e., while remaining on the hyphal cell wall, glyoxal oxidase provides hydrogen peroxide to manganese peroxidase that had diffused into the culture solution beforehand.