School of Grain Science and Technology, Jiangsu University of Science and Technology, Zhenjiang 212100, China.
School of Food Science and Technology, Jiangnan University, Wuxi 214122, China.
Molecules. 2023 Jun 18;28(12):4841. doi: 10.3390/molecules28124841.
Ochratoxin A (OTA) is the most common mycotoxin and can be found in wheat, corn and other grain products. As OTA pollution in these grain products is gaining prominence as a global issue, the demand to develop OTA detection technology has attracted increasing attention. Recently, a variety of label-free fluorescence biosensors based on aptamer have been established. However, the binding mechanisms of some aptasensors are still unclear. Herein, a label-free fluorescent aptasensor employing Thioflavin T (ThT) as donor for OTA detection was constructed based on the G-quadruplex aptamer of the OTA aptamer itself. The key binding region of aptamer was revealed by using molecular docking technology. In the absence of the OTA target, ThT fluorescent dye binds with the OTA aptamer to form an aptamer/ThT complex, and results in the fluorescence intensity being obviously enhanced. In the presence of OTA, the OTA aptamer binds to OTA because of its high affinity and specificity to form an aptamer/OTA complex, and the ThT fluorescent dye is released from the OTA aptamer into the solution. Therefore, the fluorescence intensity is significantly decreased. Molecular docking results revealed that OTA is binding to the pocket-like structure and surrounded by the A29-T3 base pair and C4, T30, G6 and G7 of the aptamer. Meanwhile, this aptasensor shows good selectivity, sensitivity and an excellent recovery rate of the wheat flour spiked experiment.
赭曲霉毒素 A（OTA）是最常见的霉菌毒素，存在于小麦、玉米和其他谷物产品中。由于这些谷物产品中的 OTA 污染作为一个全球性问题日益突出，因此开发 OTA 检测技术的需求引起了越来越多的关注。最近，已经建立了各种基于适配体的无标记荧光生物传感器。然而，一些适配体传感器的结合机制仍不清楚。在此，基于 OTA 适配体自身的 G-四链体适配体，构建了一种用于 OTA 检测的无标记荧光适配体传感器，该传感器使用噻唑橙（ThT）作为供体。通过分子对接技术揭示了适配体的关键结合区域。在不存在 OTA 靶标的情况下，ThT 荧光染料与 OTA 适配体结合形成适配体/ThT 复合物，导致荧光强度明显增强。在存在 OTA 的情况下，由于 OTA 适配体对 OTA 具有高亲和力和特异性，因此 OTA 适配体与 OTA 结合形成适配体/OTA 复合物，并且 ThT 荧光染料从 OTA 适配体释放到溶液中。因此，荧光强度显著降低。分子对接结果表明，OTA 结合到口袋状结构中，被 A29-T3 碱基对和适配体的 C4、T30、G6 和 G7 环绕。同时，该适配体传感器表现出良好的选择性、灵敏度和出色的回收率，在小麦粉加标实验中。
