Amino Acid-Programmed Biomineralization for Radical-Proximal Enzyme–MOF Interfaces in Electrochemiluminescent Influenza Hemagglutinin Sensing

Electrochemiluminescent (ECL) biosensing is fundamentally limited by inefficient coupling between catalytic radical generation and luminophore excitation. Here, we report an amino acid-programmed biomineralization strategy for constructing radical-proximal enzyme–MOF interfaces under mild aqueous conditions. Serine reconfigures zirconium precursor chemistry to convert a charge-mismatched mineralization process into a charge-adaptive one, enabling biomineralization of cationic horseradish peroxidase (HRP) within PCN-224. The resulting HRP@PCN-224(Ser) exhibits enhanced enzyme loading, preserved protein structure, improved catalytic competence, and pronounced solvent tolerance. Mechanistic analyses indicate that serine-mediated mineralization does more than facilitate enzyme incorporation: it creates a spatially integrated microenvironment in which H₂O₂ activation and ZnTCPP excitation are more effectively coupled, leading to enhanced ECL transduction. Accordingly, HRP@PCN-224(Ser) produces an approximately 9-fold stronger ECL response than the serine-free counterpart in the presence of H₂O₂. Integrated with a dual-aptamer signal-off format, this interface enables ECL sensing of influenza H1N1 hemagglutinin (HA) over 0.1–1000 ng mL^–1 with a detection limit of 0.03 ng mL^–1, together with good selectivity and satisfactory recovery in serum samples. These findings establish amino acid programmed biomineralization as an interfacial design strategy for enzyme–MOF integration and provide a mechanistically grounded framework for improving ECL bioanalysis through radical-proximal transduction.