Engineering ferritin nanocages for Cd^2 + adsorption with magnetic recyclability

Ferritin has attracted considerable attention as a biological nanoplatform due to its well-defined cage-like architecture, excellent dispersibility, and structural stability. In this study, cysteine residues were rationally introduced at the subunit interface of oyster ferritin (GKFN) to construct a thiol-enriched coordination microenvironment, aiming to enhance Cd²⁺ adsorption while reducing metal-induced protein aggregation. Biochemical characterization confirmed that the engineered ferritin (GKFN4C) maintained correct folding, preserved ferroxidase activity, and retained its self-assembled 24-mer nanocage structure. Spectroscopic analyses, dynamic light scattering (DLS), and transmission electron microscopy (TEM) revealed that the mutant ferritin exhibited improved structural stability and dispersion under high Cd²⁺ conditions compared with the wild-type protein. X-ray photoelectron spectroscopy (XPS) and isothermal titration calorimetry (ITC) further indicated that Cd²⁺ binding involved coordination interactions with amino acid residues, while the introduction of cysteine residues enhanced Cd²⁺ enrichment within the ferritin cavity. Adsorption experiments demonstrated that ferritin effectively removed Cd²⁺ from contaminated systems and maintained good structural integrity during repeated adsorption-desorption cycles. To improve the practical applicability of the system, a magnetic separation strategy was developed by introducing a His tag and employing Ni-functionalized Fe₃O₄@SiO₂ magnetic beads for protein recovery. The engineered ferritin achieved a Cd²⁺ removal efficiency of 90.7%, while the magnetic recovery strategy enabled a protein recovery efficiency of 82.6%. These findings demonstrate a recyclable ferritin-based nanobiomaterial for heavy metal removal and provide insights into the design of protein-based adsorbents.