Cellobiase activity

During purification procedures cellobiase activity was monitored by measuring nitrophenol (at A42onm) release for p-nitrophenyl-/ -D-glucoside (JO). Kinetic studies and enzyme characterization were carried out using / -D-cellobiose as substrate with the product, glucose, measured with a Beckman Glucose Analyzer (JO). Assay conditions were pH 4.8 and 50°C. 

Figure 8. Sephadex G-75 column chromatography of crude celluloses. (-O-) Protein (A 0im), (-X-) activity with respect to CMC (endoglu-canase), (- -) activity with respect to Avicel (cellobiohydrolase). Inset, top right (-O-) cellobiase activity.

The cellobiase activity in culture filtrates of T. reesei was small relative to that of cellobiohydrolase and endoglucanase. The possibility that cellobiase of T. reesei is either an intracellular or membrane-bound enzyme was indicated by experiments in which cellobiose or other carbon sources were used as the substrate for culture growth. While cellobiose can be taken up rapidly by the fungus, very little cellulase activity could be detected in the filtrate see Table IV—cellobiose as carbon source). Furthermore, the appearance of cellobiase did not parallel the appearance of cellulase activity. Increasing culture incubation time did, however, result in increasing cellobiase activity in the filtrate. This data suggested that at least some of the cellobiase present in the filtrate might have been an intracellular cellobiase which was, perhaps, released when some of... 

Carbon Source Growth0, /Dry weight, gm 200 mL culture J % Cellobiase" Activity % Cellulase Activity... 

This chapter deals with three aspects of the cellulolytic enzyme system of Thermoactinomyces sp. the location of the CM-cellulase, Avicelase, and / -glucosidase (cellobiase) activities in the culture, the multiplicity of the extracellular enzyme system, and the stability of the different activities as a function of pH, temperature, and time. The results are discussed with reference to saccharification of cellulosic materials. 

The whole culture broth and its fractions were also assayed for cellobiase activity. This activity followed closely the / -glucosidase activity, so that it was found in the culture solids and the filtrate after sonication (Table I). Only trace amounts of cellobiase activity appeared... 

Table I. / -Glucosidase and Cellobiase Activities in Whole Culture Broth, Culture Filtrate, Culture Solids, Filtrate, and Solids After Sonication for 10 Min°...

Ci activity (1,2), but / -glucosidase and cellobiase activities are probably ubiquitous (e.g., see Ref. 14 and 16). Whether or not these together constitute a cellulase complex as in fungi is unknown. The possibility seems unlikely since there are so few occasions in plants when whole cellulose microfibrils are autolyzed, and there is no need for it on nutritional grounds. Nevertheless, the question remains of why specific plant cells or tissues often elaborate more than one endocellulase. 

Whole Cells possessing Cellobiase Activity Immobilized into Hollow Fiber Membrane Reactors presented at the Europe-Japan Congress on Membranes and Membrane Processes, Stresa, Italy (1984). 

---