Endo-1, 4-β-glucanase and β-glucosidase of the ciliate Polyplastron multivesiculatum free of cellulolytic bacteria

Abstract

To determine the contribution of protozoal activity to cellulose degradation, we maintained the ciliate Polyplastron multivesiculatum free of extracellular and intracellular cellulolytic bacteria. Control experiments to verify the absence of such bacteria were performed on cellular extracts, on filtrates, and on ciliates before lyophilization. The enzymatic activity was determined by viscometry and by determining the amount of reducing sugars produced. The enzyme was found to be an endo-1, 4- β-glucanase. Polyplastron multivesiculatum which was maintained free of extracellular and intracellular bacteria degraded soluble derivatives of cellulose and slightly degraded native cellulose. The activity was not due to intracellular bacterial cellulase, as ingested bacteria were lysed and digested by proteolytic enzymes of the protozoan. In addition, when preparing the filtrate solutions, P. multivesiculatum was maintained in culture for 5 days with Streptococcus faecalis (a facultatively anaerobic, noncellulolytic bacterium). Isoelectric focusing and chromatofocusing of lyophilized P. multivesiculatum that was free of cellulolytic bacteria yielded three protein fractions that degrade carboxymethylcellulose and hydroxyethylcellulose. Chromatofocusing also revealed the presence of two protein fractions with β-glucosidase activity.