Mutant cellulases, isolation

Much effort has been expended over the years on increasing enzyme production of T. reesei by isolation of high yielding mutants and optimizing media and fermentation conditions. Strains have been isolated that produce 2-6 times the cellulase productivity of the parent wild strain (QM 6a) in batch culture. The mutants produce higher levels of cellulase protein but the specie activity of the enzymes and the proportions of the individual components (ca. 30% endo- -glucanase, 70% o- -glucanase, and less 1% cellobiase) are similar to those of the parent. 

Montenecourt and Eveleigh (37), using a special agar screening technique, have also isolated a cellulase enhanced mutant (NG-14). They suggest, however, that the cellobiohydrolase and the endoglucanase biosynthesis are regulated by different controls. They base this assumption on data for the relative ratios of cellulases obtained from this mutant needed to hydrolyze different substrates. 

More experiments are needed before any conclusions can be made. It appears that the isolation and study of a mutant which synthesizes only one cellulolytic enzyme component is needed if headway is to be made on determining the nature of regulatory control of cellulase biosynthesis. 

Selective Screening Methods for the Isolation of High Yielding Cellulase Mutants of Trichoderma reesei... 

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 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. 

Continuous culture studies for the production of cellulase have been carried out growing Trichoderma viride on a medium containing commercial D-glucose as the carbon source. Some data on cellulase biosynthesis were examined and correlated in terms of a maturation time model. Specific oxygen uptake rates were correlated with specific production rates. Mutant strains not producing cellulases have been induced and isolated from Trichoderma viride. Enrichment of the mutants was carried out by nystatin selection. 

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