Visualizing cellulase activity on the cellulose surface

Commercial exploitation of lignocellulose for biotechnological production of fuels and commodity chemicals requires efficient - usually enzymatic - saccharification of the highly recalcitrant insoluble substrate. A key characteristic of cellulose conversion is that the actual hydrolysis of the polysaccharide chains is intrinsically entangled with physical disruption of substrate morphology, which is in the spatial arrangement of the cellulose chains into solid material. This “substrate deconstruction” by cellulase activity is a slow, yet markedly dynamic process that occurs at different length scales from and above the nanometer range. Little is currently known about the role of progressive substrate deconstruction on hydrolysis efficiency. Application of advanced visualization techniques to the characterization of enzymatic degradation of different celluloses has provided important new insights, at the requisite nanoscale resolution and down to the level of single enzyme molecules, into cellulase activity on the cellulose surface.

We have prepared a mixed amorphous-crystalline cellulosic model substrate (MACS), which is shown by atomic force microscopy (AFM) to display a completely smooth surface and applied it for kinetic analysis of cellulase action. Combined kinetic and atomic force microscopy studies have been used to examine surface structural dynamics during MACS degradation. Action of the complete Trichoderma sp. cellulase system was studied, but also that of individual cellulases and auxiliary enzymes such as lytic polysaccharide monooxygenase.

Bubner P, Plank H, Nidetzky B (2013) Biotechnol Bioeng 110:1529-49

Ganner T, Bubner P, Eibinger M, Plank H, Nidetzky B (2012) J Biol Chem 287:43215-22