Thermodynamic analysis of cooperative ligand binding by the ATP-binding DNA aptamer indicates a population-shift binding mechanism

Slavkovic, Sladjana; Zhu, Yi; Churcher, Zachary R.; Shoara, Aron A.; Johnson, Anne; Johnson, Philip E.

doi:10.1038/s41598-020-76002-8

Cited by 26 publications

(30 citation statements)

References 38 publications

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“…For the set of C strands hybridizing to the 5′ region of the ATP aptamer, K D2 ATP of 5.71 ± 0.865 μM was obtained from fitting the β n dependence on K D1 ( n ) using eq S38 (Figure B, fitting coefficient of determination R 2 = 0.96). The obtained K D2 ATP value agreed well with previous reports. ,− For the set of C strands hybridizing to the 3′ region of the ATP aptamer, the K D2 ATP value of 117 ± 62.8 μM was gained (Figure C, R 2 = 0.13).…”

Section: Resultssupporting

confidence: 90%

Characterizing Aptamers with Reconfigurable Chiral Plasmonic Assemblies

Huang

Nguyen

et al. 2022

Langmuir

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Aptamers have emerged as versatile affinity ligands and as promising alternatives to protein antibodies. However, the inconsistency in the reported affinities and specificities of aptamers has greatly hindered the development of aptamer-based applications. Herein, we present a strategy to characterize aptamers by using DNA origami-based chiral plasmonic assemblies as reporters and establishing a competitive hybridization reaction-based thermodynamic model. We demonstrate the characterization of several DNA aptamers, including aptamers for small molecules and macromolecules, as well as aptamers with high and low affinities. The presented characterization scheme can be readily adapted to a wide selection of aptamers. We anticipate that our approach will advance the development of aptamer-based applications by enabling reliable and reproducible characterization of aptamers.

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Section: Resultssupporting

confidence: 90%

Characterizing Aptamers with Reconfigurable Chiral Plasmonic Assemblies

Huang

Nguyen

et al. 2022

Langmuir

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“…To confirm binding, we first used ITC. ITC is a label-free technique that measures the heat produced during binding. − When theophylline was titrated, heat release indicative of aptamer binding was observed (Figure D). By fitting the integrated heat, a dissociation constant ( K d ) of 0.51 ± 0.13 μM was obtained.…”

Section: Resultsmentioning

confidence: 99%

A DNA Aptamer for Theophylline with Ultrahigh Selectivity Reminiscent of the Classic RNA Aptamer

Huang

Liu

2022

ACS Chem. Biol.

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Since the report of the RNA aptamer for theophylline, theophylline has become a key molecule in chemical biology for designing RNA switches and riboswitches. In addition, theophylline is an important drug for treating airway diseases including asthma. The classic RNA aptamer with excellent selectivity for theophylline has been used to design biosensors, although DNA aptamers are more desirable for stability and cost considerations. In this work, we selected DNA aptamers for theophylline, and all the top sequences shared the same binding motifs. Binding was confirmed using isothermal titration calorimetry and a nuclease digestion assay, showing a dissociation constant (K d ) around 0.5 μM theophylline. The Theo2201 aptamer can be truncated down to 23-mer while still has a K d of 9.8 μM. The selectivity for theophylline over caffeine is around 250,000-fold based on a stranddisplacement assay, which was more than 20-fold higher compared to the classic RNA aptamer. For other tested analogs, the DNA aptamer also showed better selectivity. Using the structure-switching aptamer sensor design method, a detection limit of 17 nM theophylline was achieved in the selection buffer, and a detection limit of 31 nM was obtained in 10% serum.

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“…The authors found the aptamer undergoes extensive structural changes and forms a stable three-way junction upon binding. Slavkovic et al [126] used a combination of thermal stability, ITC and NMR experiments to determine why the ATP-binding aptamer binds two copies of the target cooperatively. By investigating the aptamer and two related constructs, a population shift mechanism was discovered which results in a cooperative binding mechanism.…”

Section: Nmr Spectroscopymentioning

confidence: 99%