The Use of Carbohydrate Binding Modules (CBMs) to Monitor Changes in Fragmentation and Cellulose Fiber Surface Morphology during Cellulase- and Swollenin-induced Deconstruction of Lignocellulosic Substrates

Background: Fiber fragmentation is thought to occur at dislocations, which are potential targets for the non-hydrolytic protein, Swollenin. Results: Changes in cellulose morphology within dislocations were assessed using fluorescent CBMs; Swollenin appeared to promote fragmentation at these sites. Conclusion: Swollenin targets and disrupts cellulose at fiber dislocations. Significance: Fragmentation is a key step in cellulose deconstruction and is enhanced by the actions of Swollenin.

Although the actions of many of the hydrolytic enzymes involved in cellulose hydrolysis are relatively well understood, the contributions that amorphogenesis-inducing proteins might contribute to cellulose deconstruction are still relatively undefined. Earlier work has shown that disruptive proteins, such as the non-hydrolytic non-oxidative protein Swollenin, can open up and disaggregate the less-ordered regions of lignocellulosic substrates. Within the cellulosic fraction, relatively disordered, amorphous regions known as dislocations are known to occur along the length of the fibers. It was postulated that Swollenin might act synergistically with hydrolytic enzymes to initiate biomass deconstruction within these dislocation regions. Carbohydrate binding modules (CBMs) that preferentially bind to cellulosic substructures were fluorescently labeled. They were imaged, using confocal microscopy, to assess the distribution of crystalline and amorphous cellulose at the fiber surface, as well as to track changes in surface morphology over the course of enzymatic hydrolysis and fiber fragmentation. Swollenin was shown to promote targeted disruption of the cellulosic structure at fiber dislocations.