Biomass Depolymerization projects

Surface Kinetic Mechanisms of Enzymatic Cellulose Deconstruction - Completed

Assessment of cellulose deconstruction kinetics is a critical step in the design of improved cellulases for efficient production of liquid fuels from biomass. The project elucidated molecular interactions between cellulases and cellulose at the aqueous/solid interface using surface-sensitive techniques to understand, quantitatively and mechanistically, how cellulase adsorption onto cellulose controls cleavage kinetics. Based on measured molecular-kinetic rate constants, they were able to provide molecular guides for protein engineering of improved cellulase constructs. They also developed a surface-based assay for improved enzyme development.

project Highlights

2011 Highlights

The team used their surface-based quartz crystal microbalance assay to determine Langmuir adsorption,  desorption and irreversible-binding constants for cellobiohydrolase I and endoß- glucanase, demonstrating that while CBH I adsorbs to the surface much faster, the rate of irreversible binding is identical for both enzymes.  This result suggests a binding-domain only interaction governing irreversibility. Radke’s group also explored the synergy between endoglucanase and exoglucanase on model cellulose films, finding that a high-bulk concentration of endoglucanase results in competition for surface sites and thereby slow, rather than enhanced,  cellulose degradation.

2010 Highlights

Radke’s group developed a surface-based assay to elucidate the kinetics of cellulase binding, activity, and deactivation on a solid cellulose substrate. They assessed adsorption, desorption, and denaturation rate constants for isolated endoglucanases and exoglucanases.  They then incorporated these rate constants into a novel, two-enzyme surface model for cellulase activity on a solid cellulose surface.  Cellulases adsorbed irreversibly and retained activity on the cellulose surface for only several hours after initial adsorption.

2009 Highlights

Researchers in Radke’s group began work using isolated cellulases and mixtures. As expected, endoglucanases only adsorb to the surface and do not degrade it. Exoglucanases work to degrade the surface somewhat, but it is the combination of endoglucanases and exoglucanases working in concert that is necessary to deconstruct a cellulose surface. Future work will further explore mixtures of these enzymes, pretreatment methods, and highly crystalline cellulose surfaces.

 


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