Cellulase catabolite repression

In addition to catabolite repression, the cellulase enzymes themselves are subject to end-product inhibition. For example, as glucose accumulates during saccharification, it interacts noncompetively with cellobiase to inhibit further activity of this enzyme (6). Similar inhibition of endoglucanases occurs when cellobiose accumulates in a saccharification reactor (18,19,20). 

A number of selective-screening methodologies have been devised that have allowed isolation of a series of hyperproducing and catabolite repression-resistant mutants of T. reesei. Yields of cellulase of 15 units/ mL under controlled fermentor conditions have been achieved with both Rut-NG14 and Rut-C30. Quantitative reaction of Rut-NG14 enzyme preparation with purified antibodies to cellobiohydrolase shows that in this mutant, the cellobiohydrolase is specifically hyperproduced relative to the rest of the enzymes in the cellulase complex. Rut-C30, which was derived from Rut-NG14, shows resistance to catabolite repression for... 

The relationship between catabolite repression by glucose and induction of cellulase by sophorose has been studied in T. viride by Nisizawa and co-workers (36, 37). The induction by sophorose (10 M) was competitively repressed by glucose and other metabolites such as pyruvate. Since glucose was an effective repressor when added one hour after the previous addition of actinomycin D, it was concluded that the repression takes place at the translational level. Previous work indicated (26) that the sophorose induction led to the formation of a cellulase component designated FII, which is the source of cellulase II discussed below. In higher plants indoleacetic acid (38) and abscisic acid (39) have been shown to stimulate cellulase production. 

Pseudomonas fluorescens produced two extracellular (A and B) and one cell-bound (C) cellulase components, the latter being released by treatment with EDTA-lysozyme in isotonic sucrose. Culture with 0.5% glucose formed little cellulase. Cellobiose stimulated only the synthesis of C. The formation of A and B was strikingly enhanced in cultures with cellulose, sophorose, or continuous low concentration of cellobiose. The absence of extracellular cellulase synthesis in 0.5% cellobiose culture may be caused by catabolite repression. The three cellulases were purified and characterized. None of them split cellobiose, but all hydrolyzed various cellodextrins and celluloses. C easily attacked cellotriose and cellotriosyl sorbitol, but A and B had no effect. When pure B was incubated with broken spheroplasts of sophorose-grown cells, a cellulase component indistinguishable from A was formed. 

Since the inhibition of cellulase synthesis in 0.5% cellobiose culture seems to be caused by so-called catabolite repression as reported for the formation of many other hydrolases, the same results would be expected for other metabolizable sugars, which will show an enhanced cellulase synthesis depending on the supply conditions. Most of the sugars tested... 

In defined mixed culture the substrate is pasteurized or steriUzed and inoculated simultaneously with more than one pure culture. This can be beneficial for complex substrates and where the various strains use different carbon sources. For example, mixed cultures of Trichoderma reesei or Chaetomium cel-lulolyticum with Candida lipolytica resulted in increased protein production from wheat straw because the yeast uses glucose and prevents catabolite repression of the fungal cellulase [66,67]. 

The development of an agar plate screening technique has allowed the isolation of a range of mutants of Trichoderma reesei capable of biosynthesizing cellulase under conditions of high catabolite repression. Commercial aspects of the production and isolation of hyper-cellulase-producing mutants of T. reesei were discussed. 

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