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Metal-Mediated DNA Adsorption on Carboxylated, Hydroxylated and Hydrogenated Nanodiamonds
Federated Research Data Repository / dépôt fédéré de données de recherche
Zandieh, Mohamad; Liu, Juewen — 2023-08-17 Nanodiamonds (NDs) have attracted considerable attention owing to their quantum properties and versatility in biological applications. In this study, we systematically investigated the adsorption of DNA oligonucleotides onto NDs with three types of surface groups: carboxylated (COOH-), hydroxylated (OH-), and hydrogenated (H-). Among them, only the H-NDs showed fluorescence quenching property that is useful for real-time DNA adsorption kinetic studies. The effect of common metal ions on DNA adsorption was studied. In the presence of Na+, the order of DNA adsorption efficiency was H-> OH- > COOH-, whereas all the NDs showed a similar DNA adsorption efficiency in the presence of divalent metal ions such as Ca2+ and Zn2+. Desorption studies revealed that hydrogen bonding and metal-mediated interactions were dominant for the adsorption of DNA, and the H-NDs exhibited extraordinarily tight DNA adsorption. Finally, a fluorescently labeled DNA was adsorbed on NDs for DNA detection, and the COOH-NDs had the highest target specificity, and a detection limit of 1.4 nM was achieved. This study indicates the feasibility of using metal ions to mediate the physical adsorption of DNA to NDs and compares various NDs with graphene oxide for fundamental understanding.
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Electrostatic-mediated Binding of DNA to Lysozyme: Evaluation of Aptamer-based Assays for Highly Positively Charged Targets
Federated Research Data Repository / dépôt fédéré de données de recherche
Shi, Lu; Jin, Yan; Liu, Juewen — 2023-10-04 Lysozyme is a highly popular protein target for the development of aptamer-based biosensors. Because lysozyme is a polycation and DNA is a polyanion, it is essential to separate the contribution of nonspecific electrostatic interactions from specific aptamer binding. In this study, various factors affecting the binding of DNA and lysozyme, including DNA sequence, DNA length, pH, and salt concentration, were explored using fluorescence polarization. We concluded that direct fluorescence polarization and fluorescence intensity change are unlikely to be directly applicable for aptamer-based biosensors to detect lysozyme because all tested DNA sequences showed binding. These fundamental studies confirm the dominant role of electrostatic binding. We further evaluated three other methods including label-free fluorescent detection using a DNA staining dye, label-free colorimetric detection using gold nanoparticles, and a fluorescent sensor based on the strand displacement reaction. In each case, we focused on a random DNA sequence that is not expected to bind to lysozyme as an aptamer. Of all the methods, only the strand displacement strategy can be potentially used to evaluate aptamer binding, as the other methods all responded to non-aptamer sequences. This study provides valuable insights for assaying aptamer binding to cationic proteins that can exhibit nonspecific attraction to DNA.
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Cross-Binding of Adenosine by Aptamers Selected Using Theophylline
Federated Research Data Repository / dépôt fédéré de données de recherche
Huang, Po-Jung Jimmy; Li, Albert Zehan; Dieckmann, Thorsten; Liu, Juewen — 2023-10-20 We recently reported that some adenosine binding aptamers can also bind caffeine and theophylline with around 20-fold lower affinities. This discovery led to the current work to examine the cross-binding of adenosine to theophylline aptamers. For the DNA aptamer for theophylline, cross-binding to adenosine was observed, and the affinity was 18 to 38-fold lower for adenosine based on assays using isothermal titration calorimetry and ThT fluorescence spectroscopy. The binding complexes were characterized using NMR spectroscopy, and both adenosine and theophylline showed an overall similar binding structure to the DNA theophylline aptamer, although small differences were also observed. In contrast, the RNA aptamer did not show binding to adenosine, although both aptamers have very similar relative selectivity for various methylxanthines including caffeine. After a negative selection, a few new aptamers with completely different primary sequences for theophylline were obtained and they did not show binding to adenosine. Thus, there are many ways for aptamers to bind theophylline and some can have cross-binding to adenosine. In biology, theophylline, caffeine, and adenosine can bind to the same protein receptors to regulate sleep, and their binding to the same DNA motifs may suggest an early role of nucleic acids in similar regulatory functions.

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(auteur : Liu, Juewen)

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