| dc.description.abstract | The aim of this study is to address the interaction issues of enzyme immobilization within metal-organic frameworks (MOFs). Although the innovative in-situ synthesis method (de novo mild water-based) developed by the laboratory in 2015 successfully encapsulated enzymes, the interactions between the enzyme and the MOF affected enzyme activity. To improve the interaction between materials and enzymes, hollow material synthesis methods (Hollow MOFs, HMOFs) were developed. However, the etching process used to prepare hollow materials, along with the environmental conditions and released substances like MOF linkers, still impacted enzyme activity. Therefore, optimizing etching conditions to maintain the enzyme’s original activity is a critical issue.
In this study, different MOF materials were selected to prepare hollow structures, combined with the ball milling method developed by our laboratory in 2023. This method is ultra-fast (10 seconds), environmentally friendly, and high-yield (about 80%). Catalase (CAT), UiO-66 (Universitetet i Oslo-66), and ZIF-90 (Zeolitic Imidazole Framework-90) were successfully used to synthesize a hierarchical MOF material, CAT-on-UiO-66@ZIF-90. By utilizing the differential resistance of the materials in buffer to the etching solution, the inner layer material, UiO-66, was selectively etched while the stable outer layer material, ZIF-90, was retained. This process successfully formed the hollow material CAT@HZIF-90.
Characterization measurements confirmed the successful synthesis of the hollow structure. Activity tests showed that the enzyme’s freedom within the hollow material was enhanced, resulting in a significant increase in activity. The activity constant increased from 3.5 × 10-2 s-1 for the solid material to 7.2 × 10-2 s-1 for the hollow material, indicating a twofold increase in activity. Compared to HZIF-8, which had an activity of 1.1 × 10-2 s-1, HZIF-90 demonstrated a sevenfold increase in activity. Subsequent immersion of the material in a proteinase-K solution further confirmed that the enzyme was primarily encapsulated within the MOF.
Finally, for future applications, the combination of fluorescence spectroscopy and hollow materials could be used to study enzyme unfolding and refolding processes, observing the dynamic changes in enzyme structure. The unfolding and refolding of the enzyme structure within the hollow material were explored. After treatment with urea and subsequent removal, the refolding of the enzyme structure was evident. This demonstrates that the HZIF-90 system has significant advantages in stabilizing enzyme structures, providing a new option for future enzyme immobilization. | en_US |