The mechanism of enzymatic cellulose degradation. Purification and some properties of two different 1,4beta-glucan glucanohydrolases from Trichoderma viride.
1. A low-molecular-weight and a high-molecular-weight 1,4beta-glucan glucanohydrolase (Cx enzyme) have been isolated from a commercial cellulase preparation derived from culture filtrates of the fungus Trichoderma viride. 2. The purification method for the isolation of the low-molecular-weight enzyme is a three-step procedure including chromatography on Bio-Gel P-10, chromatography on a dipolar adsorbent (arginine-Sepharose 6 B) and isoelectric focusing. 3. The starting material for the isolation of the high-molecular-weight enzyme was pre-fractionated by chromatography on Bio-Gel P-10, by DEAE-Sephadex chromatography and by SE-Sephadex chromatography as described previously by us. Further fractionation of this material was achieved by affinity chromatography and repeated isoelectric focusing. 4. Free zone electrophoresis of the low-molecular-weight enzyme indicated a homogeneous protein. The high-molecular-weight enzyme was homogenous in sedimentation equilibrium analysis. 5. The molecular weights of the enzymes were 12 500 and 50 000 +/- 2000 respectively. The former value was determined by chromatography on a calibrated column of Bio-Gel P-100 and the latter value by sedimentation equilibrium analysis. 6. The low-molecular-weight enzyme was isoelectric at pH 4.60 (10 degrees C) and contained 21% carbohydrate. The corresponding values for the high-molecular-weight enzyme were pH 3.39 and 12%. 7. Both enzymes were active in releasing free fibers from filter-paper. The low-molecular-weight enzyme was estimated to be about twice as effective as the high-molecular-weight enzyme in this regard.
D000818:Animals; D002480:Cellulase / Q000302:isolation & purification / Q000378:metabolism*; D002482:Cellulose / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000378:metabolism*; D006601:Hexoses / Q000032:analysis; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism*; D046911:Macromolecular Substances; D003904:Mitosporic Fungi / Q000201:enzymology*; D008970:Molecular Weight; D010209:Paper; D014242:Trichoderma / Q000201:enzymology*
D006601:Hexoses; D007527:Isoenzymes; D046911:Macromolecular Substances; D002482:Cellulose; D006026:Glycoside Hydrolases; D002480:Cellulase