Dataset Information

Isolation of Natural DNA Aptamers for Challenging Small-Molecule Targets, Cannabinoids.

ABSTRACT: Aptamers are nucleic acid-based affinity reagents that are isolated via an in vitro 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 (-)-trans-?⁹-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.

SUBMITTER: Yu H

PROVIDER: S-EPMC7895305 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Isolation of Natural DNA Aptamers for Challenging Small-Molecule Targets, Cannabinoids.

Yu Haixiang H Luo Yingping Y Alkhamis Obtin O Canoura Juan J Yu Boyang B Xiao Yi Y

Analytical chemistry 20210202 6

Aptamers are nucleic acid-based affinity reagents that are isolated via an <i>in vitro</i> 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 ...[more]

PMID: 33528997

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Project description:Applications of synthetic biology spanning human health, industrial bioproduction, and ecosystem monitoring often require small molecule sensing capabilities, typically in the form of genetically encoded small molecule biosensors. Critical to the deployment of greater numbers of these systems are methods that support the rapid development of such biosensors against a broad range of small molecule targets. Here, we use a previously developed method for selection of RNA biosensors against unmodified small molecules (DRIVER) to perform a selection against a densely multiplexed mixture of small molecules, representative of those employed in high-throughput drug screening. Using a mixture of 5,120 target compounds randomly sampled from a large diversity drug screening library, we performed a 95-round selection and then analyzed the enriched RNA biosensor library using next generation sequencing (NGS). From our analysis, we identified RNA biosensors with at least 2-fold change in signal in the presence of at least 217 distinct target compounds with sensitivities down to 25 nM. Although many of these biosensors respond to multiple targets, clustering analysis indicated at least 150 different small-molecule sensing patterns. We also built a classifier that was able to predict whether the biosensors would respond to a new compound with an average precision of 0.82. Since the target compound library was designed to be representative of larger diversity compound libraries, we expect that the described approach can be used with similar compound libraries to identify aptamers against other small molecules with a similar success rate. The new RNA biosensors (or their component aptamers) described in this work can be further optimized and used in applications such as biosensing, gene control, or enzyme evolution. In addition, the data presented here provide an expanded compendium of new RNA aptamers compared to the 82 small molecule RNA aptamers published in the literature, allowing further bioinformatic analyses of the general classes of small molecules for which RNA aptamers can be found.

Dataset Information

Isolation of Natural DNA Aptamers for Challenging Small-Molecule Targets, Cannabinoids.

Publications

Isolation of Natural DNA Aptamers for Challenging Small-Molecule Targets, Cannabinoids.

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