Endotoxin contamination, as one of the most significant challenges in recombinant protein production by Escherichia coli, represents a critical biosafety concern and greatly hinders the biomedical application of recombinant proteins. Conventional methods, such as extreme-condition inactivation and chromatography-based separation, are plagued by issues including protein denaturation, low efficiency, and operational complexity in endotoxin removal. In this work, we developed a novel magnetic bifunctional endotoxin removal nano-agent (MagBER) with a multi-layered structure, consisting of a superparamagnetic FeO core, a mesoporous TiO intermediary layer, and an outer shell functionalized with boronic acid groups. This dual-functional design significantly enhances endotoxin removal efficiency through the TiO layer and boronic acid groups, ensuring stable endotoxin clearance performance even in high-salt environments and complex biological matrices. MagBER exhibits reusability while maintaining protein structural integrity. Moreover, MagBER has been successfully employed for endotoxin removal in various proteins, establishing it as a promising and sustainable solution for endotoxin clearance in biopharmaceutical applications.