16-04: Molecular-level mechanisms of enzymatic deconstruction of cellulose

We present recent insights from molecular simulations of the enzymatic deconstruction of cellulose. In particular, we have examined the three sub-domains of the Family 7 cellulase (Cel7A) from Hypocrea jecorina to understand the structure-function relationships of each sub-domain alone. We have investigated the behavior of the carbohydrate-binding module (CBM) on cellulose at multiple resolutions and calculated the binding affinity of various CBM mutants for direct comparison to experiment. The flexibility of the Cel7A linker has been examined with and without glycosylation with respect to other linkers from fungal cellulases, which demonstrate significant homology differences. We have conducted multiple thermodynamic mutations of aromatic residues in the catalytic tunnel, which likely affect processivity, and examined the thermodynamics of product inhibition in the Cel7A wild type enzyme and various mutants. For cellulose, we have completed a large-scale comparison of commonly used force fields for cellulose Iβ, and calculated the work that a processive enzyme must conduct to depolymerize cellulose chains for four polymorphs of cellulose. These insights on individual components of cellulase action on cellulose have been assimilated into recent simulations of the entire enzyme acting on cellulose, which have been extended to the sub-microsecond time scale to obtain a realistic version of the Cel7A catalytically-active complex and to study processivity of various Cel7A mutants. Overall, these results provide new details regarding the molecular-level behavior of processive cellulases.