14-32: Addition of a carbohydrate-binding module enhances cellulase penetration into cellulose substrates

Cellulases are of great interest for application in biomass degradation, yet the molecular details of the mode of action of glycoside hydrolases during degradation of insoluble cellulose remains elusive. To further improve these enzymes for application at industrial conditions, it is critical to gain a better understanding of not only the details of the degradation process, but also the function of accessory modules. Toward that end, we fused a cellulose-binding module (CBM) from family 2a to two thermophilic endocellulases. Catalytic activity of the chimeric enzymes was enhanced up to nine fold on insoluble cellulose substrates as compared to wild type. We then applied neutron reflectometry to determine the mechanism of these enhancements. Importantly, we demonstrate that the wild type enzymes affect primarily the surface properties of an amorphous cellulose film, while the chimeras containing a CBM alter the bulk properties of the amorphous film. These findings suggest that the CBM2a improves the efficiency of these cellulases by enabling digestion of cellulose located in the bulk of the film that could not be digested by the endocellulases not containing the CBM.