One-step, reagent-free construction of nano-enzyme as visual and reusable biosensor for oxidase substrates.
One-step, reagent-free construction of nano-enzyme as visual and reusable biosensor for oxidase substrates.
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DOI：
10.1016/j.aca.2023.342008
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发表时间：
2023-11
影响因子：
6.2
通讯作者：
Ke Quan;Yuqing Zeng;Wenke Zhang;Fengfeng Li;Mengjiao Li;Zhihe Qing;Linlin Wu
中科院分区：
文献类型：
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作者：
Ke Quan;Yuqing Zeng;Wenke Zhang;Fengfeng Li;Mengjiao Li;Zhihe Qing;Linlin Wu
酶的底物是与人体生理健康密切相关的生物必需物质。然而，目前的生物传感方法由于受到杂质的干扰，存在难以回收利用和酶易变性的问题。在此，我们以葡萄糖氧化酶(GOx)为模型酶，研制了一种可重复使用的可视化底物生物传感器。GOX在−20℃下一步固定到金纳米颗粒(AuNPs)上，不需要额外的试剂。由此产生的纳米酶通过与以TMB为底物的辣根过氧化物酶(HRP)偶联来产生生化信号。结果表明，固定化GOX对葡萄糖检测具有令人满意的灵敏度(0.68UμM)，在恶劣条件下比游离GOX具有更高的固有稳定性，能够可靠地检测复杂液体(有色饮料和唾液)中的葡萄糖。此外，纳米酶在催化氧化4个循环后仍保持80%的活性。这一策略构建了一种通用的纳米酶底物生物传感器，它只依赖于酶体内的固有半胱氨酸，而避免了功能性的手柄修饰。
The substrates of oxidase are biologically essential substances that are closely associated with human physiological health. However, current biosensing methods suffer from tough recyclability and undesired denaturation of enzyme due to impurity interference. Herein, we have developed a visual and reusable biosensor for detecting substrate using glucose oxidase (GOx) as a model oxidase. GOx was immobilized onto gold nanoparticles (AuNPs) at −20 °C in one step without additional reagents. The resulting nano-enzyme generated coloimetric signals by coupling with horseradish peroxidase (HRP) using TMB as the substrate. Our results demonstrated that the immobilized GOx exhibited satisfactory sensitivity (0.68 μM) for glucose detection and higher inherent stability than free GOx under harsh conditions, enabling reliable detection of glucose in complex fluids (colored beverages and saliva). Furthermore, the nano-enzyme retained 80 % activity even after four cycles of catalytic oxidation. This strategy constructs a universal biosensor for substrates with nano-enzyme which rely only on intrinsic cysteine within the oxidase while avoiding functional handle modification.