(192d) Mechanistic Insights Into Cellulose III Formation and Its Depolymerization

Authors: 
Chundawat, S. P., Michigan State University
Uppugundla, N., Michigan State University
Gao, D., Michigan State University
Balan, V., Great Lakes Bioenergy Center, Michigan State University
Dale, B., Michigan State University
Bellesia, G., T6 and CNLS - Los Alamos National Laboratory
Langan, P., Biosciences Division - Los Alamos National Laboratory
Gnanakaran, S., T6 - Los Alamos National Laboratory


Enzymatic conversion of cellulose can be improved via either reducing cellulose crystallinity by thermochemical treatments or enhancing cellulase activity by protein engineering. A third approach is via reorganizing the hydrogen bonding network within crystalline cellulose to produce a less recalcitrant allomorph. Cellulose III is one such allomorph, however, there is little known about its formation and depolymerization by glycosyl hydrolases. Detailed studies were carried out to determine the effect of ammonia-water-acetone loading, total residence time and reaction temperature on the conversion of cellulose I to III. Cellulose crystallinity and extent of conversion between I and III allomorphic states was determined using X-ray diffraction (spectral deconvolution approach). The substrates were then subjected to hydrolysis by various combinations of purified Trichoderma reesei cellulases. Based on protein structural analysis and activity assay data, it appears that a cellulase such as Cel7B with a more open and unrestricted active site cleft may further enhance the degradation of cellulose III. Molecular dynamics simulations also revealed that cellulose III fibrils are readily hydrated, explaining its reduced hydrophobically-driven binding to cellulases and that their surface has structural and dynamical features that significantly influences binding of cellulases to it. Glucan chains within cellulose III are more readily accessible by cellulases due to reduced intra-chain and intra-sheet hydrogen bonding compared to native cellulose.