Linking Protein Encapsulation in ZIF-8 with Its Biofunction: Molecular Dynamics Simulation Validated by Experiments

Correlating encapsulated protein with its bioactivity remains challenging, particularly for bone morphogenetic protein-2 (BMP-2), where both high loading efficiency and bioactivity must be ensured. In the study, molecular dynamics (MD) simulation was applied to elucidate the encapsulation process of BMP-2 within ZIF-8 and validated its functional consequences by experimental characterization in vitro. MD simulations suggest that BMP-2 rapidly anchors to the main crystal facets ((011), (112), and (222)) of ZIF-8 through predominant coordination bonds, accounting for over 57% of the adsorption energy. This facet-specific anchoring may guide subsequent framework growth, leading to confined encapsulation of the protein within the ZIF-8 lattice with a more compact yet structurally rigid conformation. Experimentally, the as-synthesized BMP-2@ZIF-8 features intraparticle nanodomains with a radius of gyration of 22.3 ± 3.2 Å, confirming single-protein confinement, and achieved an encapsulation efficiency of approximately 89%. Upon release, BMP-2 retained its native secondary structure and exhibited significantly enhanced osteogenic differentiation of bone marrow mesenchymal stem cells compared to free BMP-2, as evidenced by elevated alkaline phosphatase activity, calcium nodule formation, and activation of the p-Smad1/5/8 signaling pathway. A “facet-anchored guided growth” pathway, distinct from classical protein-induced biomineralization, was proposed to link the protein encapsulation process with its function. This work provides insight into the fundamental gap between encapsulated protein stability and its biofunction, facilitating the rational design of therapeutic protein delivery systems.