Purification and characterization of a crystalline cellulose hydrolyzing enzyme of tricholderma harzianum

ABSTRACT

An Avicel-hydrolyzing cellulase was isolated from a shake flask culture of Trichoderma harzianum and purified by Sephadex G-75 gel filtration and DEAE-Sephadex A-50 chromatography. This cellulose component, called cellulase F, showed a single protein band by SDS-PAGE. Its molecular weight was found to be 50, 100 by SDS-PAGE and 47, 800 by gel filtration. It was most active at pH 4.5 to 5.0 and a temperature of 50°C. The enzyme was stable at the pH range 4.0 to 5.5 and lost its activity rapidly beyond the temperature of 50°C. Cellulose F was rich in acidic amino acids but poor in basic and sulphur containing amino acids. The hydrolytic activity of cellulase F on Avicel, filter paper and cotton was higher as compared to those on the soluble substrates. The hydrolytic products of the insoluble substrates contained high levels of cellobiose and cellotriose but low levels of glucose. Cellulase F was an essential component of the cellulase complex since it showed strong synergism with other components of the complex