Activation of Lysozyme with Robust Flavoenzyme Activity by Amyloid Fibrillation

The intrinsic disadvantages of natural enzymes limit their practical applications. How to achieve artificial enzymes with a green process, low cost, high stability, robust catalytic activity, and excellent biocompatibility remains a great challenge. Amyloid fibrils deserve particular attention in electron transfer and biocatalysis due to their unique biochemical properties and long-range ordered structure. Inspired by the relationship between the structure and function of natural flavoenzymes, we report that lysozyme amyloid fibril (LAF) binds cofactor flavin mononucleotide (FMN) and forms a stable nanofibril complex (^FMNLAF) through noncovalent interactions. Interestingly, ^FMNLAF exhibits NADH oxidase activity (K_m, 55.3 μM; K_cat, 0.65 min^-1) to generate NAD⁺ under physiological conditions. Possessing exceptional structural stability, ^FMNLAF shows enhanced activity at high temperature (60 °C) and tolerates organic solvents. In addition, ^FMNLAF also exhibits strong Fe³⁺-cyt c reductase-like activity in aerated environments. This work provides a convenient and rational strategy for designing metal-free biocatalytic systems through amyloid fibrillation, which converts a protein into an artificial enzyme.