CONTINUOUS MONITORING OF CELLULOSE OXIDATION BY CELLOBIOSE OXIDASE FROM PHANEROCHAETE-CHRYSOSPORIUM

Cellobiose oxidase activity from Phanerochaete chrysosporium was monitored continuously by two techniques that are independent of the presence of a solid substrate, measuring either ferricyanide by the electrode potential of anaerobic ferricyanide plus ferrocyanide, or O2 with a specific electrode. Both methods demonstrated that microcrystalline cellulose (Avicel) is a substrate for oxidation. All experiments were at pH 4.0 and 30-degrees-C. Rates were evaluated for two-electron transfer. For Avicel --> ferricyanide, a study of initial rates gave k(cat) = 0.40 +/- 0.05 s-1, much slower than k(cat) = 3.6 +/- 0.3 s-1 with cellobiose as substrate. With Avicel plus 0.2-mu-M cellobiose oxidase, the rate fell to half its initial value in 3-8 min, indicating a heterogeneity of reducing ends. For Avicel --> O2, k(cat) = 0.057 +/- 0.004 s-1. Half maximal rates corresponded to K(m)(Avicel) = 8 +/- 2 g l-1 with ferricyanide, and 3.3 +/- 0.5 g l-1 with O2 as acceptor.