Harnessing fragmented cells as biocatalysts: The critical size for sustaining the integrity of the electron transport chain

Abstract Biocatalysis has emerged as a cornerstone of sustainable manufacturing, yet conventional modes are hindered by inherent limitations such as metabolic interference, mass transfer barriers, and instability. This study presented a novel platform using fragmented Gluconobacter oxydans , combining the autonomous cofactor regeneration of whole cells with the superior substrate accessibility of free enzymes. It was observed that subcellular membrane fragments retain dehydrogenase activity and an intact electron transport chain (ETC), with a critical size threshold (37,300–250,000 g centrifugal force) systematically validated for sustaining this function. The fragmented cell system eliminates carbon diversion by decoupling catalysis from central metabolism, achieving near‐complete substrate conversion across multiple dehydrogenase substrates. Furthermore, artificial electron transfer experiments confirmed the essential role of ETC coupling in the catalytic mechanism. A fully functional FCM system could serve as a scalable and efficient biocatalytic tool for industrial bioconversion processes.