Department of Chemistry and Biochemistry, Florida International University, 11200 SW 8th Street, Miami, Florida, 33199, United States.
State Key Laboratory of Natural Medicines, Jiangsu Key Laboratory of TCM Evaluation and Translational Research, Department of Complex Prescription of TCM, China Pharmaceutical University, Nanjing 211198, P. R. China.
Anal Chem. 2021 Feb 16;93(6):3172-3180. doi: 10.1021/acs.analchem.0c04592. Epub 2021 Feb 2.
Aptamers are nucleic acid-based affinity reagents that are isolated via an process known as systematic evolution of ligands by exponential enrichment (SELEX). Despite their great potential for a wide range of analytical applications, there are relatively few high-quality small-molecule binding aptamers, especially for "challenging" targets that have low water solubility and/or limited moieties for aptamer recognition. The use of libraries containing chemically modified bases may improve the outcome of some SELEX experiments, but this approach is costly and yields inconsistent results. Here, we demonstrate that a thoughtfully designed SELEX procedure with natural DNA libraries can isolate aptamers with high affinity and specificity for challenging small molecules, including targets for which such selections have previously failed. We first isolate a DNA aptamer with nanomolar affinity and high specificity for (-)--Δ-tetrahydrocannabinol (THC), a target previously thought to be unsuitable for SELEX with natural DNA libraries. We subsequently isolate aptamers that exhibit high affinity and cross-reactivity to two other challenging targets, synthetic cannabinoids UR-144 and XLR-11, while maintaining excellent specificity against a wide range of non-target interferents. Our findings demonstrate that natural nucleic acid libraries can yield high-quality aptamers for small-molecule targets, and we outline a robust workflow for isolating other such aptamers in future selection efforts.
适体是通过一种称为指数富集的配体系统进化（SELEX）的过程分离得到的基于核酸的亲和试剂。尽管它们在广泛的分析应用中有很大的潜力，但相对较少有高质量的小分子结合适体，特别是对于水溶性低和/或适体识别有限的“具有挑战性”的靶标。使用含有化学修饰碱基的文库可能会改善一些 SELEX 实验的结果，但这种方法成本高，并且结果不一致。在这里，我们证明了使用天然 DNA 文库精心设计的 SELEX 程序可以分离出对具有挑战性的小分子具有高亲和力和特异性的适体，包括以前认为不适合使用天然 DNA 文库进行 SELEX 的靶标。我们首先分离出一种 DNA 适体，对 (-)--Δ-四氢大麻酚（THC）具有纳摩尔亲和力和高特异性，THC 是以前认为不适合使用天然 DNA 文库进行 SELEX 的靶标。随后，我们分离出对两种其他具有挑战性的靶标，合成大麻素 UR-144 和 XLR-11，具有高亲和力和交叉反应性的适体，同时对广泛的非靶标干扰物保持出色的特异性。我们的研究结果表明，天然核酸文库可以为小分子靶标产生高质量的适体，并且我们概述了在未来的选择工作中分离其他此类适体的稳健工作流程。
