Abstract
In many respects, enzymes offer advantages over traditional chemical processes due to their decreased energy requirements for function and inherent greener processing. However, significant barriers exist for the utilization of enzymes in industrial processes due to their limited stabilities and inability to operate over larger temperature and pH ranges. Immobilization of enzymes onto solid supports has gained attention as an alternative to traditional chemical processes due to enhanced enzymatic performance and stability. This study demonstrates the co-immobilization of glucose oxidase (GOx) and horseradish peroxidase (HRP) as an enzyme system on Metal-Organic Frameworks (MOFs), UiO-66 and UiO-66-NH2, that produces a more effective biocatalyst as shown by the oxidation of pyrogallol. The two MOFs utilized as solid supports for immobilization were chosen to investigate how modifications of the MOF linker affect stability at the enzyme/MOF interface and subsequent activity of the enzyme system. The enzymes work in concert with activation of HRP through the addition of glucose as a substrate for GOx. Enzyme immobilization and leaching studies showed HRP/GOx@UiO-66-NH2 immobilized 6% more than HRP/GOx@UiO-66, and leached only 36% of the immobilized enzymes over three days in the solution. The enzyme/MOF composites also showed increased enzyme activity in comparison with the free enzyme system: the composite HRP/GOx@UiO-66-NH2 displayed 189 U/mg activity and HRP/GOx@UiO-66 showed 143 U/mg while the free enzyme showed 100 U/mg enzyme activity. This increase in stability and activity is due to the amine group of the MOF linker in HRP/GOx@UiO-66-NH2 enhancing electrostatic interactions at the enzyme/MOF interface, thereby producing the most stable biocatalyst material in solution. The HRP/GOx@UiO-66-NH2 also showed long-term stability in the solid state for over a month at room temperature.
Highlights
Due to current advances in biotechnology, the commercial value of enzymes as biocatalysts has increased dramatically in recent years [1,2,3,4]
Based on our results for both UiO-66 and UiO-66-NH2, as well as the results reported by Lou et al, it is apparent that Metal-Organic Frameworks (MOFs) linker functionality plays a crucial role in maximizing loading and catalytic activity by enzyme immobilization on MOF supports
Bio-rad Dye Protein Assay Dye Reagent Concentrate, and Bio-rad Lyophilized Bovine γ-Globin was used to form a standard concentration calibration curve. The results of this investigation indicate that horseradish peroxidase (HRP)/glucose oxidase (GOx) @UiO-66-NH2 is the better biocatalyst for pyrogallol oxidation by activity and stability measurements
Summary
Due to current advances in biotechnology, the commercial value of enzymes as biocatalysts has increased dramatically in recent years [1,2,3,4]. The use of biocatalysts for industrial applications is attractive because of the benefits offered by enzyme proteins, including high selectivities, diverse functionalities, and the promotion of greener chemistries [4]. One strategy of preparing enzymes that are more robust to withstand pH, temperature, and/or organic solvents is by immobilizing enzymes on solid supports. In this way, enzymes may be altered that may increase their activity, specificity, or selectivity of the target reaction [5]. There is a great deal of Catalysts 2020, 10, 499; doi:10.3390/catal10050499 www.mdpi.com/journal/catalysts
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