Understanding Free and Complexed Enzyme Mechanisms and Factors Contributing to Cell Wall Recalcitrance
Tuesday, April 29, 2014: 1:00 PM
Grand Ballroom F-G, lobby level (Hilton Clearwater Beach)
Michael Resch1, Bryon Donohoe1, Peter N. Ciesielski1, Ashutosh Mittal1, Rui Katahira2, Eric Karp2, Michael E. Himmel1, Stephen Decker1 and Gregg T. Beckham2, (1)Biosciences Center, National Renewable Energy Laboratory, Golden, CO, (2)National Bioenergy Center, National Renewable Energy Laboratory, Golden, CO
Fungal free enzymes and bacterial complexed cellulosomes deconstruct biomass using different physical mechanisms. Free enzymes, which typically contain a large proportion of GH7 cellobiohydrolase, diffuse throughout the substrate and hydrolyze primarily from the cellulose reducing end, resulting in “sharpened” macrofibrils. In contrast, complexed cellulosomes contain a diverse array of carbohydrate binding modules and multiple catalytic specificities leading to delamination and physical peeling of the cellulose macrofibril structures. To investigate how cellulose structure contributes to recalcitrance, we compared the deconstruction of cellulose I, II, and III-b using free and complexed enzyme systems. We also evaluated both systems on Clean Fractionation and alkaline pretreated biomass, which remove much of the lignin, to determine the impact on enzyme loading reduction. Free fungal enzymes demonstrated a swelling of the outer surface of the plant cell walls while removing localized disruptions, resulting in a smooth surface appearance. Cellulosomes produced cell wall surfaces with localized areas of disruption and little surface layer swelling.  These studies contribute to the overall understanding of biomass recalcitrance and how combining different enzymatic paradigms may lead to the formulation of new enzyme cocktails to reduce the cost of producing sugars from plant cell wall carbohydrates.