1-08: Heterologous expression of cellulases in the filmentous fungus Neurospora crassa

Currently the enzyme cost contribution to bioethanol generation from cellulosic biomass is a significant limiting factor to producing a cost-competitive fuel. The development of a more efficient enzyme or enzyme production system will reduce this cost. Filamentous fungi have the capacity to produce large amounts of proteins, and this quality has been exploited by industry to produce specific proteins in quantities exceeding 40 g/L. These high titers, however, are achieved only with homologous protein production. In order for the use of engineered enzymes and enzyme cocktails to be practical, these proteins must be produced in quantities similar to homologous fungal proteins. Our research is directed towards assessing the ability of the filamentous fungus Neurospora crassa to express heterologous cellulases and improving production of both individual cellulases and cellulase cocktails. In this study, we evaluate the expression of native cellobiohydrolase I (CBHI) of N. crassa and heterologous CBHIs inserted in the resident locus of N. crassa. The transcriptome and secretome of wildtype N. crassa was characterized and compared with that of heterologous CBHI knock-in strains. The activities of the secretomes were also assessed.