(10c) Exploration of Significant Enzyme Activities Required for the Deconstruction of Ionic-Liquid Pretreated Biomass

Terrestrial biomass is a second generation renewable feedstock, comprised of polysaccharides derived from cellulose and hemicellulose, which can be used to produce carbon-based chemicals and fuels.  The recalcitrance to deconstruction of biomass is one of the main impediments in the conversion of biomass to monomeric sugars and the production of sugar platform fuels and chemicals.  Ionic Liquid (IL) pretreatment creates unique biomass substrates composed of amorphous cellulose with largely intact hemicellulose and lignin. IL pretreated substrates differ from many other pretreatments which remove hemicellulose and retain cellulose crystallinity. This substrate poses an interesting challenge for enzymatic deconstruction using traditional commercial mixtures.

Mono-component cellulase and hemicellulase enzymes from Aspergillus nidulans have been expressed, purified, and characterized. Key enzyme activities and/or interactions that significantly contribute to the conversion of a variety of lignocellulosic biomass substrates to monomeric sugars were examined by enzyme combinations in series of hydrolysis experiments.  Auxiliary hemicellulases seem to be essential to the mono-component enzyme mixtures.  Investigation of these accessory enzymes also yields insight into an understanding of the differences in hemicellulose structure between feedstocks.  Some auxiliary hemicellulases include acetyl xylan esterases (AXE), arabinofuranosidases, and α-glucuronidases.  In addition to mono-component enzyme mixtures, next generation commercially-available enzyme mixtures were characterized and formulated using key activities to optimize hydrolysis at low enzyme loadings.  These simplified mixtures will be dependent on the substrate.  As part of these studies, an analysis of variance (ANOVA) was used to statistically model the main and interaction effects of key enzyme activities for each substrate.