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Salt-mediated two-site ligand binding by the cocaine-binding aptamer

Multisite ligand binding by proteins is commonly utilized in the regulation of biological systems and exploited in a range of biochemical technologies. Aptamers, although widely utilized in many rationally designed biochemical systems, are rarely capable of multisite ligand binding. The cocaine-bind...

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Autores principales: Neves, Miguel A.D., Slavkovic, Sladjana, Churcher, Zachary R., Johnson, Philip E.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5388413/
https://www.ncbi.nlm.nih.gov/pubmed/28025391
http://dx.doi.org/10.1093/nar/gkw1294
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author Neves, Miguel A.D.
Slavkovic, Sladjana
Churcher, Zachary R.
Johnson, Philip E.
author_facet Neves, Miguel A.D.
Slavkovic, Sladjana
Churcher, Zachary R.
Johnson, Philip E.
author_sort Neves, Miguel A.D.
collection PubMed
description Multisite ligand binding by proteins is commonly utilized in the regulation of biological systems and exploited in a range of biochemical technologies. Aptamers, although widely utilized in many rationally designed biochemical systems, are rarely capable of multisite ligand binding. The cocaine-binding aptamer is often used for studying and developing sensor and aptamer-based technologies. Here, we use isothermal titration calorimetry (ITC) and NMR spectroscopy to demonstrate that the cocaine-binding aptamer switches from one-site to two-site ligand binding, dependent on NaCl concentration. The high-affinity site functions at all buffer conditions studied, the low-affinity site only at low NaCl concentrations. ITC experiments show the two ligand-binding sites operate independently of one another with different affinities and enthalpies. NMR spectroscopy shows the second binding site is located in stem 2 near the three-way junction. This ability to control ligand binding at the second site by adjusting the concentration of NaCl is rare among aptamers and may prove a useful in biotechnology applications. This work also demonstrates that in vitro selected biomolecules can have functions as complex as those found in nature.
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spelling pubmed-53884132017-04-18 Salt-mediated two-site ligand binding by the cocaine-binding aptamer Neves, Miguel A.D. Slavkovic, Sladjana Churcher, Zachary R. Johnson, Philip E. Nucleic Acids Res Chemical Biology and Nucleic Acid Chemistry Multisite ligand binding by proteins is commonly utilized in the regulation of biological systems and exploited in a range of biochemical technologies. Aptamers, although widely utilized in many rationally designed biochemical systems, are rarely capable of multisite ligand binding. The cocaine-binding aptamer is often used for studying and developing sensor and aptamer-based technologies. Here, we use isothermal titration calorimetry (ITC) and NMR spectroscopy to demonstrate that the cocaine-binding aptamer switches from one-site to two-site ligand binding, dependent on NaCl concentration. The high-affinity site functions at all buffer conditions studied, the low-affinity site only at low NaCl concentrations. ITC experiments show the two ligand-binding sites operate independently of one another with different affinities and enthalpies. NMR spectroscopy shows the second binding site is located in stem 2 near the three-way junction. This ability to control ligand binding at the second site by adjusting the concentration of NaCl is rare among aptamers and may prove a useful in biotechnology applications. This work also demonstrates that in vitro selected biomolecules can have functions as complex as those found in nature. Oxford University Press 2017-02-17 2016-12-26 /pmc/articles/PMC5388413/ /pubmed/28025391 http://dx.doi.org/10.1093/nar/gkw1294 Text en © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Chemical Biology and Nucleic Acid Chemistry
Neves, Miguel A.D.
Slavkovic, Sladjana
Churcher, Zachary R.
Johnson, Philip E.
Salt-mediated two-site ligand binding by the cocaine-binding aptamer
title Salt-mediated two-site ligand binding by the cocaine-binding aptamer
title_full Salt-mediated two-site ligand binding by the cocaine-binding aptamer
title_fullStr Salt-mediated two-site ligand binding by the cocaine-binding aptamer
title_full_unstemmed Salt-mediated two-site ligand binding by the cocaine-binding aptamer
title_short Salt-mediated two-site ligand binding by the cocaine-binding aptamer
title_sort salt-mediated two-site ligand binding by the cocaine-binding aptamer
topic Chemical Biology and Nucleic Acid Chemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5388413/
https://www.ncbi.nlm.nih.gov/pubmed/28025391
http://dx.doi.org/10.1093/nar/gkw1294
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