A single multifunctional enzyme mediates a sequential three-step biocascade

Traditional biocatalytic cascades typically require discrete enzymes for each synthetic step. Here, we report unprecedented trifunctional imine reductases (IRED) that conduct three sequential transformations—alkene reduction, intramolecular reductive amination, and imine reduction—all within a single catalytic cycle. This elegant single-enzyme catalytic system directly transforms linear substrates into enantiomerically pure 2-aryl pyrrolidines via a concerted cascade without intermediate isolation. Combining density functional theory (DFT) calculations and mechanistic studies, we elucidate how the IRED achieves step-selective catalysis. Our findings establish a proof-of-concept for simplifying complex biocascades using multifunctional enzymes, offering a powerful strategy to streamline synthetic pathways.