5-45: Investigating the expression, secretion and enzymatic activities of the cellulolytic machinery of the filamentous ascomycete fungus, Neurospora crassa

The model filamentous ascomycete fungus, Neurospora crassa, can be easily isolated in nature from the stems of tropical grasses and has the capability to degrade both lignin and cellulose. Our lab has used transcriptional profiling combined with proteomics research to characterize proteins/enzymes involved in plant cell wall degradation. To further understand how fungal enzymes attack the substrate and how they work synergistically to degrade the components of Miscanthus cell walls, we generated two versions of vectors to epitope-tag candidate genes that encode proteins identified by mass spec analysis of secreted N. crassa proteins. These constructs will be used for investigating the expression, secretion, and enzymatic activities of cellulases induced by cellulose using the native promoter and constructs under the regulation of a promoter expressed in minimal medium in N. crassa. Biochemical subtraction experiments will be used to evaluate changes in biochemical activity of depleted enzyme mixtures to identify known protein function and perhaps unknown proteins that have crucial roles in cellulose and plant cell wall degradation.