Biocatalytic Upcycling of Biomass via Immobilized Enzymes in Continuous Flow

Abstract Agroindustrial waste streams such as sugar beet pulp (SBP) are rich in underutilized uronic acids, representing an abundant yet valuable source of renewable carbon. Here, we report a modular biocatalytic system using cellulose-immobilized enzyme cascades under continuous flow to produce polymer precursors at the gram scale from waste-derived feedstocks. Key to this process is the fusion of uronate dehydrogenase and ene reductase enzymes with engineered cellulose-binding domains (CBDs), allowing efficient co-immobilization on cellulose supports. These enzymes achieved >95% conversion of uronic acids and retained activity over eight continuous flow cycles. The system operates under mild aqueous conditions and integrates in-line cofactor recycling and product extraction, enabling the production of building blocks in up to 97% isolated yields. The resulting diacid and lactone products were further transformed into (poly)esters and (poly)amides with potential applications in bioplastics, surfactants, and functional materials. This modular, flow-compatible biocatalytic strategy provides a generalizable route for valorizing uronic acid-rich waste streams, advancing strategies for biomanufacturing.