Carbohydrate recognition mechanisms of glycoside hydrolase modular domains

Enzymatic hydrolysis of biomass is often conducted with multi-modular cellulase enzymes consisting at least one catalytic domain and one or more carbohydrate binding module (CBM) domains. Here, we present recent molecular modeling efforts focused on defining the mechanisms by which these domains recognize their carbohydrate substrates. First, we examine cello-oligomer binding within two Family 4 CBMs using molecular dynamics simulations and ligand binding free energy calculations. Binding site dynamics contribute to observed variation in specificity between the two CBMs from the same Family. Additionally, we investigate the thermodynamic feasibility of hypothesized bi-directional cello-oligomer binding to the CBM, which has implications for how these CBMs assist in targeting non-crystalline cellulose deconstruction. Second, we investigate the carbohydrate recognition mechanisms of glycoside hydrolase catalytic domains using a Family 18 chitinase model system. We use free energy calculations and molecular simulation alongside experimental measurements of activity and processivity to uncover the roles of individual aromatic residues within the active sites of two processive chitinases. The results from this latter study represent a significant step forward in our efforts to characterize a general structure-function relationship in processive glycoside hydrolases, which are critical components of industrial biomass conversion cocktails.