Domain structure and function of α-1,3-glucanase from Bacillus circulans KA-304, an enzyme essential for degrading basidiomycete cell walls.

Bacillus circulans KA-304 α-1,3-glucanase (Agl-KA) includes an N-terminal discoidin domain (DS1), a carbohydrate binding module family 6 (CB6), threonine and proline repeats (TPs), a second discoidin domain (DS2), an uncharacterized conserved domain (UCD), and a C-terminal catalytic domain. Domain deletion enzymes lacking DS1, CB6, and DS2 exhibited lower α-1,3-glucan-hydrolyzing and -binding activities than the wild type, Agl-KA. An α-1,3-glucan binding assay with fluorescent protein fusion proteins indicated that DS1, CB6, and DS2 bound to α-1,3-glucan and fungal cell walls, and that binding efficiency was increased by their combined action. In contrast, UCD did not exhibit any α-1,3-glucan-binding activity. A dramatic decrease in protoplast formation in the Schizophyllum commune mycelium was observed given only a DS1 deletion. An Agl-KA with deletion DS1, CB6, and DS2 produced no protoplasts. These results indicate that the combined actions of DS1, CB6, and DS2 contributed to increased cell-wall binding and were indispensable for efficient Agl-KA cell-wall degradation.