Chitooligosaccharide oxidase (ChitO) is a fungal carbohydrate oxidase containing a bicovalently bound FAD cofactor. The enzyme is known to catalyse the oxidation of chitooligosaccharides, oligomers of N-acetylated glucosamines derived from chitin degradation. In this study, the unique substrate acceptance was explored by testing a range of N-acetyl-D-glucosamine derivatives, revealing that ChitO preferentially accepts carbohydrates with a hydrophobic group attached to C2. The enzyme also accepts streptozotocin, a natural product used to treat tumours. Elucidation of the crystal structure provides an explanation for the high affinity towards C2-decorated glucosamines: the active site has a secondary binding pocket that accommodates groups attached at C2. Docking simulations are fully in line with the observed substrate preference. This work expands the knowledge on this versatile enzyme.
- covalent flavin
- crystal structure