An extracellular enzyme which utilizes molecular oxygen to oxidize cellodextrins to the corresponding aldonic acids has been isolated from culture filtrates of the white-rot fungus Sporotrichum pulverulcntum. This enzyme, tentatively named cellobiose oxidase, has been highly purified by classical techniques and has been demonstrated to be a glycoprotein with a molecular weight of approximately 93000. Ultraviolet spectra of the enzyme in the presence and absence of substrate are characteristic of a hemoprotein. Acidic hydrolyses of the enzyme followed by a spectrofluorimetric investigation of the hydrolysate has demonstrated the presence of approximately one flavin component per enzyme molecule. The possible role of this complex enzyme in cellulose degradation is discussed.The white-rot fungus Sporotrichum pulverulentum is capable of degrading both the polysaccharides and the lignin present in wood. The extracellular enzyme systems utilized by the fungus for the degradation of cellulose have been studied extensively in our kdboratory. A recent review article [l] summarizes the present state of knowledge.For the degradation of cellulose the fungus utilizes the following hydrolytic enzymes, namely; five endo-1,4-fi-glucanases, one exo-l,4-b-glucanase and one or several P-glucosidases. The endo-glucanases and the exo-glucanase have been purified and characterized and a synergistic action between these enzymes has been demonstrated 12 -51. The fi-glucosidases are presently being studied. In addition to the hydrolytic enzymes acting on cellulose or cellodextrins, S. pulverulentum produces an extracellular enzyme, cellobiose : quinone oxidoreductase, of importance in cellulose as well as in lignin degradation. Its function is to reduce phenoxy radicals and quinones formed by the action of phenol oxidases on degradation products from the lignin, thereby oxidizing cellobiose and higher cellodextrins to the corresponding lactones The present information on the enzymes involved in cellulose degradation has permitted a partial reconstruction of the extracellular enzyme system of S. pufverulentum. Preliminary experiments with such a reconstructed system indicate that the presence of the known hydrolytic enzymes is not sufficient to account [6 -81.Enzymes. Endo-1.4-P-glucanase (EC 3.2.1.4): P-glucosidase (EC 3.2.1.21).for the cellulose degrading activity of culture filtrates [9]. These studies indicate that the missing component requires molecular oxygen to stimulate cellulose degradation.A search for oxidative enzymes involved in cellulose degradation by S. pulverulentum has resulted in the purification and partial characterization of an extracellular hemoprotein enzyme (different from the previously studied cellobiose : quinone oxidoreductase) which oxidizes cellobiose and cellodextrins using molecular oxygen. A description of this oxidase, given the tentative name of cellobiose oxidase, and a discussion of its role in cellulose degradation is the subject of this paper. MATERIALS AND METHODS The OrganismThe fungus used in ...

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In order to find naturally oeeurring white-rot fungi which preferentially degrade lignin, 25 different species of such fungi were cultivated on pine wood blocks and on kraft lignin agar plates with and without cellulose. Due to differenees in phenol oxidase reactions on the kraft lignin agar plates, the 25 fungi could be divided into two groups, 1 and 2, which also differed in other properties. The three Group 1 fungi Sporotrichum pulverulentum, Phanerochaete sp. L-l and Polyporus dichrous produced high levels of endo-l,4-/?-glucanase and eeliobiose :quinone oxidoreductase in shaking cellulose fiasks and a low level of phenol oxidase in standing wood meal flasks. The four fungi Merulius tremellosus, Phlebia radiata, Pycnoporus cinnabarinus and Pleurotus ostreatus from Group 2, on the other hand, produeed low levels of endo-1,4-/S-glucanase and eeliobiose :quinone oxidoreduetase in the cellulose flasks and a high level of phenol oxidase in the wood meal fiasks.Analyses of pine wood blocks degraded by the above-mentioned fungi in the presence of either malt extract, asparagine or NH4H2PO4 revealed that malt extract gave good lignin degradation.In the presence of this nutrient source, P, cinnabarinus, at 3.4% weight loss, even degraded 12.5% lignin without loss of cellulose or mannan. No common degradation pattern was, however, obtained using malt extract, asparagine or NH^HjPOj. It is suggested that white-rot fungi, which preferentially degrade lignin, may be found among Group 2 fungi producing large amounts of phenol oxidases.