AbstractAmong the extensive repertoire of carbohydrate-active enzymes, lytic polysaccharide monooxygenases (LPMOs) have a key role in recalcitrant biomass degradation. LPMOs are copper-dependent enzymes that catalyze oxidative cleavage of glycosidic bonds in polysaccharides such as cellulose and chitin. Several LPMOs contain carbohydrate-binding modules (CBMs) that are known to promote LPMO efficiency. Still, structural and functional properties of some of these CBMs remain unknown and it is not clear why some LPMOs, like CjLPMO10A from Cellvibrio japonicus, have two CBMs (CjCBM5 and CjCBM73). Here, we studied substrate binding by these two CBMs to shine light on the functional variation, and determined the solution structures of both by NMR, which includes the first structure of a member of the CBM73 family. Chitin-binding experiments and molecular dynamics simulations showed that, while both CBMs bind crystalline chitin with Kd values in the µM range, CjCBM73 has higher affinity than CjCBM5. Furthermore, NMR titration experiments showed that CjCBM5 binds soluble chitohexaose, whereas no binding to soluble chitin was detected for CjCBM73. These functional differences correlated with distinctly different architectures of the substrate-binding surfaces of the two CBMs. Taken together, these results provide insight into natural variation among related chitin-binding CBMs and the possible functional implications of such variation.
Substrate Binding Effects of Carbohydrate Binding Modules on the Catalytic Activity of a Multifunctional Cellulase
