Localization of Cellobiose Dehydrogenase in Cellulose-Grown Cultures ofPhanerochaete chrysosporium

Abstract

To elucidate the function of cellobiose dehydrogenase (CDH) in cellulose degradation byPhanerochaete chrysosporium,production and localization of CDH were investigated and compared with those in shaking and aerated static cultures grown on cellulose. Substantial CDH activity was detected in the medium of the shake cultures after 8 days of incubation, while no CDH activity was detected in the medium of static cultures at any point during the incubation period. Light microscopy clearly showed that many cellulose particles were adsorbed on the surface of the hypha in static cultures, whereas no cellulose particles were adsorbed to the hypha in shake cultures. The addition of laminarinase to static cultures was very effective in detaching cellulose particles from the hypha surfaces. Using a potentiometric assay performed with an oxidation–reduction potential electrode, some CDH activity could be detected on the hypha/cellulose complexes in static cultures. Thus, CDH is produced also in static cultures, albeit in lower amounts than in shake cultures, but the enzyme is not released into the medium. It seems likely that the β-1,3-glucan layer plays an important role in CDH localization and cellulose degradation. Immunocytochemical confocal laser scanning microscopy for the static cultures demonstrated that most CDH was adsorbed on the surface of the cellulose, especially around the cracks, which were formed by the action of cellulases during the course of incubation. From these observations, we conclude a direct participation of CDH in the degradation of cellulose in cooperation with cellulases.