Modular assembly of dual-template artificial antibody-antigen-directed co-immobilization of bioreactor for in-situ synthesis of epoxy compounds: Leveraging oxidized nicotinamide adenine dinucleotide cofactor to enhance yield.

Enzymatic epoxidation offers eco-friendly advantages over chemical methods. This work developed a modular dual-enzyme reactor through artificial antibody-antigen directed immobilization for in-situ epoxide biosynthesis. Magnetic artificial antibodies were synthesized using hydroquinone and 4-aminopyrimidine templates, while 2,5-dihydroxybenzaldehyde and 4-aminopyrimidine-5-carbaldehyde were employed to modify glucose oxidase (GOx) and lipase (LIP), respectively, forming artificial antigens. The dual-enzyme reactor was constructed through affinity recognition between artificial antibodies and antigens, a modular strategy that allowed rational regulation of enzyme ratios in the co-immobilized system to match cascade reaction requirements, thereby promoting optimal yield. This GOx-LIP dual-enzyme reactor achieved alkene epoxidation with epoxide yields exceeding 70% under optimized conditions. Notably, the yield could be further enhanced to 90% by incorporating oxidized nicotinamide adenine dinucleotide as a cofactor. This modular biological platform demonstrates the superiority of multi-enzyme cascade catalysis in alkene epoxidation.