(222a) The Relationship Between Chitinase Binding Affinity, Processivity, and Work Required to Decrystallize β-Chitin

Authors: 
Jana, S., University of Kentucky
Payne, C. M., National Renewable Energy Laboratory
Hamre, A. G., Norwegian University of Life Sciences
Sørlie, M., Norwegian University of Life Sciences
Structural polysaccharides, such as cellulose and chitin, are highly crystalline in nature owing to their complex networks of intra- and inter-molecular hydrogen bonds; this provides strong resistance to microbial and chemical attack. Nature evolves enzyme cocktails, including processive glycoside hydrolases (GHs), to hydrolyze these recalcitrant polysaccharides to soluble sugars. Between subsequent hydrolytic reactions, processive GHs remain attached to the polymeric substrate, and hence, are responsible for the majority of hydrolytic bond cleavage. However, our understanding of the relationship between processive enzyme function and the crystalline polymorph and morphology is limited. We hypothesize that the free energy of binding an oligosaccharide to the active site of a processive GH is directly related to the ability of an enzyme to processively hydrolyze a polymer chain from a crystal; moreover, the ligand binding free energy of a processive enzyme must also be more thermodynamically favorable than the work required to decrystallize the polymer. We have selected the Serratia marcescens Family 18 chitinase model system, including processive chitinases, ChiA and ChiB and a non-processive chitinase, ChiC, to test our hypothesis. Combining molecular dynamics simulations and free energy calculations, we find that processive ChiA and ChiB exhibit ligand binding free energies that are more thermodynamically favorable than the work to decrystallize a chito-oligosaccharide from the crystalline β-Chitin surface, which is essential for forward processive movement. The non-processive ChiC binds chito-oligosaccharides with a free energy that is significantly less favorable than the work of decrystallization. In general, our findings suggest that processive GH function necessitates tight binding within the enzyme active site. Comparisons to cellulases aid in understanding the transferability of these findings to other processive GHs.