Characterization of endo-1,4-beta-D-glucanases purified from Trichoderma viride

Abstract

Four electrophoretically distinct endo-1,4-beta-D-glucanases (EC 3.2.1.4) from Trichoderma viride have been identified and named as isozymes, Endoglucanases I, II, III and IV, according to their electrophoretic mobilities on polyacrylamide gels. Endoglucanases II, III and IV, the homogeneity of each of which was established by discontinuous gel electrophoresis and ultracentrifugation, had specific activities on CM-cellulose of 1010, 60 and 250 specific fluidity units/mg protein, respectively. These enzymes have similar pH optima (pH 4.0-4.5) and are labile at pH values greater than 8.0. The endoglucanases are high in acidic and hydroxylated amino acids and glycine, but low in basic amino acids. Values of 12.0, 10.3 and 13.1 have been determined for the epsilon 1%280 of purified Endoglucanases II, III and IV, respectively. Sedimentation equilibrium analysis has established the molecular weights of Endoglucanases II, III and IV to be 37 200, 52 000 and 49 500, respectively. The three endoglucanases contain mannose, galactose, glucose and glucosamine. Mannose is the principal neutral sugar in each enzyme. Endoglucanase II is distinguished by its low carbohydrate content, 4.5% (w/w), compared to Endoglucanases III and IV which contain 15.0% and 15.2% carbohydrate, respectively.