2,3‐Dihydroxybenzoic Acid Decarboxylase from Fusarium oxysporum: Crystal Structures and Substrate Recognition Mechanism

A 2,3‐dihydroxybenzoic acid decarboxylase from Fusarium oxysporum (2,3‐DHBD_Fo) has a relatively high catalytic efficiency for the decarboxylation of 2,3‐dihydroxybenzoic acid (DHBA) and carboxylation of catechol, thus it has a different substrate spectrum from other benzoic acid decarboxylases. We have determined the structures of 2,3‐DHBD_Fo in its apo form and complexes with catechol or 2,5‐dihydroxybenzoic acid at 1.55, 1.97, and 2.45 Å resolution, respectively. The crystal structures of 2,3‐DHBD_Fo show that the enzyme exists as a homotetramer, and each active center has a Zn2+ ion coordinated by E8, H167, D291 and three water molecules. This is different from 2,6‐DHBD from Rhizobium sporomusa, in which the Zn2+ ion is also coordinated with H10. Surprisingly, mutation of A10 of 2,3‐DHBD_Fo to His resulted in almost complete loss of the enzyme activity. Enzyme‐substrate docking and site‐directed mutation studies indicate that residue R233Δ interacts with the 3‐hydroxy group of 2,3‐DHBA, and plays an important role in substrate recognition for this enzyme, thus revealing the molecular basis 2,3‐dihydroxybenzoic acid decarboxylase. It depends on your position: Crystal structure and mutation studies reveal that R233Δ plays an important role in the substrate recognition of 2,3‐dihydroxybenzoic acid decarboxylase (DHBD) from Fusarium oxysporum, as well as the molecular basis for this enzyme being a 2,3‐DHBD with different substrate preference from known 2,6‐DHBDs and salicylic acid decarboxylases.