Purification and characterization of cellobiose dehydrogenases from the white rot fungus Trametes versicolor

• Published in: Applied and Environmental Microbiology Vol. 62; no. 12; pp. 4417 - 4427
• Main Authors: Roy, B.P. (Air Products and Chemicals Inc., Allentown, PA.), Dumonceaux, T, Koukoulas, A.A, Archibald, F.S
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01.12.1996
• Subjects: ACTIVIDAD ENZIMATICA; ACTIVITE ENZYMATIQUE; ALCOHOL DESHIDROGENASA; ALCOOL DESHYDROGENASE; AZUCARES; Biological and medical sciences; Biology of microorganisms of confirmed or potential industrial interest; Biotechnology; CATALASA; CATALASE; CATION; CATIONES; CELLOBIOSE; CELLULOSE; CELOBIOSA; CELULOSA; COBRE; COENZIMAS; COENZYME; Copper; CORIOLUS VERSICOLOR; CUIVRE; FER; Fundamental and applied biological sciences. Psychology; Fungi; HIERRO; Iron; LACASA; LACCASE; MANGANESE; MANGANESO; METAL; METALES; Microbiology; Mission oriented research; OXIDACION; OXYDATION; PASTA; PATE CELLULOSIQUE; PESO MOLECULAR; Physiology and metabolism; POIDS MOLECULAIRE; PURIFICACION; PURIFICATION; QUINONAS; QUINONE; REDUCCION; REDUCTION; SECUENCIA NUCLEOTIDICA; SEQUENCE NUCLEOTIDIQUE; SINTESIS DE PROTEINAS; SUCRES; SYNTHESE PROTEIQUE; TEMPERATURA; TEMPERATURE; Trametes versicolor; Purification; Enzyme; Industrial application; Coriolus versicolor; Basidiomycetes; Environmental factor; Dehydrogenase; Paper industry; Fungi; Characterization; Enzymatic activity; Substrate specificity; Cellobiose dehydrogenase (quinone); Oxidoreductases; Kinetic parameter; Physicochemical properties; Thallophyta
• ISSN: 0099-2240; 1098-5336
• DOI: 10.1128/aem.62.12.4417-4427.1996

The white rot fungus Trametes versicolor degrades lignocellulosic material at least in part by oxidizing the lignin via a number of secreted oxidative and peroxidative enzymes. An extracellular reductive enzyme, cellobiose dehydrogenase (CDH), oxidizes cellobiose and reduces insoluble Mn(IV)O2, commonly found as dark deposits in decaying wood, to form Mn(III), a powerful lignin-oxidizing agent. CDH also reduces ortho-quinones and produces sugar acids which can promote manganese peroxidase and therefore ligninolytic activity. To better understand the role of CDH in lignin degradation, proteins exhibiting cellobiose-dependent quinone-reducing activity were isolated and purified from cultures of T. versicolor. Two distinct proteins were isolated; the proteins had apparent molecular weights of 97,000 and 81,000 and isoelectric points of 4.2 and 6.4, respectively. The larger CDH (CDH 4.2) contained both flavin and heme cofactors, whereas the smaller contained only a flavin (CDH 6.4). These CDH enzymes were rapidly reduced by cellobiose and lactose and somewhat more slowly by cellulose and certain cello-oligosaccharides. Both glycoproteins were able to reduce a very wide range of quinones and organic radical species but differed in their ability to reduce metal ion complexes. Temperature and pH optima for CDH 4.2 were affected by the reduced substrate. Although CDH 4.2 showed rather high substrate specificity among the ortho-quinones, it could also rapidly reduce a structurally very diverse collection of other species, from negatively charged triiodide ions to positively charged hexaquo ferric ions. CDH 6.4 showed a higher Km and a lower Vmax and turnover number than did CDH 4.2 for all substrates tested. Furthermore, CDH 6.4 did not reduce the transition metals Fe(III), Cu(II), and Mn(III) at concentrations likely to be physiologically relevant, while CDH 4.2 was able to rapidly reduce even very low concentrations of these ions. The reduction of Fe(III) and Cu(II) by CDH