This study presents the engineering of a bacterial eugenol oxidase to selectively dehydrogenate 4-n-propylguaiacol, a lignin-derived compound, into isoeugenol, a valuable flavor and fragrance molecule. Through computational predictions and a multi-step engineering process, the researchers improved the enzyme's thermostability, chemoselectivity, and activity. Structural insights revealed that slow catalytic activity was due to a slowly decaying covalent substrate-flavin adduct, which was addressed by targeted mutations. The final engineered variant, PROGO, exhibited good activity, thermostability, and high chemoselectivity for 4-n-propylguaiacol oxidation. Gram-scale preparative reactions demonstrated PROGO's utility as a stable biocatalyst for isoeugenol production from lignin.

Yiming Guo, Laura Alvigini, Milos Trajkovic, Lur Alonso-Cotchico, Emanuele Monza, Simone Savino, Ivana Marić, Andrea Mattevi, Marco W. Fraaije