Magnetically Recoverable δ-FeOOH Particles for Multilayer Enzyme Immobilization and Surface-Induced Activity Tuning

Enzyme immobilization is an effective strategy to enhance enzyme stability, which remains a major drawback in biocatalysis. However, achieving high enzyme loading without loss of activity is still a key limitation of conventional support. Most supports that overcome these limitations, such as nano- and microparticles, involve high production costs. Therefore, a support that combines the advantages of high surface area, easy recovery, and low-cost production is still lacking in the literature. In this study, we present a low-cost, dual-function, magnetically recoverable support material, superparamagnetic δ-FeOOH (feroxyhyte) particles, that enhances enzyme activity and modulates protein structure both prior to, and following immobilization. We show that δ-FeOOH enables lipase immobilization at loadings exceeding 60 mg g^-1 while maintaining activity and structural integrity. Circular dichroism and fluorescence analyses reveal support-induced conformational changes, decreased α-helicity and increased β-sheet content, that do not impair enzymatic performance. Zeta potential analysis further confirms progressive surface saturation and multilayer formation, with continued adsorption beyond ∼40 mg g^-1, without functional decline. Notably, both lipase and NADH oxidase (L<i>p</i>NOX) exhibit up to 1.3-fold activity enhancement in the presence of δ-FeOOH, even in the absence of covalent binding, suggesting a surface-induced activation mechanism. Together, these findings establish δ-FeOOH as a high-capacity, structurally tunable enzyme support. Its ability to promote both immobilization and functional enhancement makes it a promising platform for next-generation biocatalysts in continuous, high-density, and multienzyme systems.