Department of Chemistry, Faculty of Science, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.
Institute of Nanoscience and Nanotechnology (ION2), Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.
Sci Rep. 2024 Oct 11;14(1):23859. doi: 10.1038/s41598-024-72503-y.
An alternative biomarker for assessing the cyanide levels in postmortem materials is crucial for investigating acute cyanide intoxication. Herein, an aptamer-ligand biorecognition system with high specificity was developed to detect acute cyanide poisoning via its secondary metabolite, 2-amino-2-thiazoline-4-carboxylic acid (ATCA). Potential aptamers were screened from a random library of 66-base single-stranded DNA using GO-SELEX, with individual aptamers being identified through single-stranded DNA sequencing. Molecular docking was employed to predict the affinity of these aptamers toward ATCA and selected counter-targets; these predictions were confirmed using thermodynamic analysis with an isothermal titration calorimeter. Owing to its label-free biomolecular binding interactions, Apt46 exhibited the highest affinity against ATCA and notable selectivity against structurally similar counter-targets. Thus, an amino-tagged Apt46 binding aptamer was attached to a carbon electrode modified with EDC-NHS-activated graphene oxide. The binding of Apt46 to ATCA was quantified by measuring current changes using differential pulse voltammetry. The aptasensor achieved a detection limit of 0.05 µg/mL and demonstrated suitability for detecting ATCA across various biological matrices, with the high recovery percentages ranging from 92.29 to 114.22%. Overall, the proposed ATCA aptasensor is promising for identifying ATCA metabolites in cases of acute cyanide exposure.
一种用于评估死后材料中氰化物水平的替代生物标志物对于研究急性氰化物中毒至关重要。在此，开发了一种具有高特异性的适体-配体生物识别系统，通过其次级代谢产物 2-氨基-2-噻唑啉-4-羧酸（ATCA）来检测急性氰化物中毒。通过 GO-SELEX 从 66 碱基单链 DNA 随机文库中筛选出潜在的适体，通过单链 DNA 测序鉴定出单个适体。分子对接用于预测这些适体与 ATCA 和选定的对照靶标之间的亲和力；这些预测通过使用等温滴定量热仪进行的热力学分析得到了证实。由于其无标记的生物分子结合相互作用，Apt46 对 ATCA 表现出最高的亲和力和对结构相似的对照靶标的显著选择性。因此，将氨基标记的 Apt46 结合适体连接到用 EDC-NHS 激活的氧化石墨烯修饰的碳电极上。通过使用差分脉冲伏安法测量电流变化来定量测定 Apt46 与 ATCA 的结合。该适体传感器的检测限为 0.05μg/mL，并且在各种生物基质中都显示出适合检测 ATCA 的能力，回收率高，范围为 92.29%至 114.22%。总的来说，所提出的 ATCA 适体传感器有望用于鉴定急性氰化物暴露情况下的 ATCA 代谢物。
