Engineering Neurospora Crassa for Improved Cellobionate Production from Cellulose

The conventional gluconic acid and isobutanol production process involves five steps: pretreatment, cellulase production, enzymatic hydrolysis to make sugars, aerobic fermentation to produce gluconic acid from cellulose hydrolysate, and anaerobic fermentation to produce isobutanol from sugars. Cellulase production or purchase represents a substantial portion of processing costs. A new route for converting cellulosic biomass to isobutanol and gluconate was proposed. In this proposed route, cellulase production, enzymatic hydrolysis, and aerobic fermentation will be consolidated into a single aerobic fermentation step where cellooligosaccharide aldonates are produced from pretreated cellulose by an engineered cellulolytic fungus, Neurospora crassa.  Isobutanol and gluconate will be produced from the cellooligosaccharide aldonates in a subsequent anaerobic fermentation step.   In this study, we report engineering N. crassa for high yield cellobionate production from cellulose, including fermentation optimization and kinetic analysis of cellulose degradation.