Partial purification, characterization and regulation of cellulolytic enzymes from <i>Trichoderma longibrachiatum</i>

Abstract

A net purification of 9·46‐, 18·6‐ and 16·7‐fold for filter paper (FP) hydrolytic activity, carboxymethyl (CM) cellulase and β‐glucosidase, respectively was achieved through ion exchange and gel chromatographies. The purified enzyme preparation showed an optimal pH of 5·0 for CM cellulase and 5·5 for the other two components. The enzyme activities increased up to 60°–65°C for the three enzyme components and they were stable at 30° or 40°C and pH 4·5 to 5·0 after 20–30 min treatment. The four enzyme components, that is, two FP activities (unadsorbed and adsorbed), a CM cellulase and a β‐glucosidase, had K m values of 47·6 mg, 33·3 mg, 4·0 mg and 0·18 mmol/l with V max of 4, 1·28, 66·5 and 1·28 units per mg protein. The molecular weights as determined with SDS‐PAGE were found to be 44000, 38000, 55000 and 63000 for the above four enzyme components in the same sequence. A distinct type of synergistic action was observed between these components by their action on dewaxed cotton. Glycerol at 1% strongly repressed the formation of all the cellulolytic enzymes. The role of proteolytic enzymes in in vitro inactivation of cellulases was not apparent.