Reduction in Dynamics of Base pair Opening upon Ligand Binding by the Cocaine-Binding Aptamer
| Autor: | Devid Garaev, Philip E. Johnson, Howard N. Hunter, Zachary R. Churcher |
|---|---|
| Rok vydání: | 2020 |
| Předmět: |
0303 health sciences
Base pair Aptamer Dynamics (mechanics) Biophysics Articles Aptamers Nucleotide Ligands 03 medical and health sciences Crystallography chemistry.chemical_compound 0302 clinical medicine Cocaine chemistry Nucleic Acid Conformation Magnetization transfer Binding site Base Pairing Protein secondary structure 030217 neurology & neurosurgery DNA 030304 developmental biology Cocaine binding |
| Zdroj: | Biophys J |
| ISSN: | 0006-3495 |
| DOI: | 10.1016/j.bpj.2020.08.012 |
| Popis: | We have used magnetization transfer NMR experiments to measure the exchange rate constant (kex) of the imino protons in the unbound, cocaine-bound, and quinine-bound forms of the cocaine-binding DNA aptamer. Both long-stem 1 (MN4) and short-stem 1 (MN19) variants were analyzed, corresponding to structures with a prefolded secondary structure and ligand-induced-folding versions of this aptamer, respectively. The kex values were measured as a function of temperature from 5 to 45°C to determine the thermodynamics of the base pair opening for MN4. We find that the base pairs close to the ligand-binding site become stronger upon ligand binding, whereas those located away from the binding site do not strengthen. With the buffer conditions used in this study, we observe imino 1H signals in MN19 not previously seen, which leads us to conclude that in the free form, both stem 2 and parts of stem 3 are formed and that the base pairs in stem 1 become structured or more rigid upon binding. This is consistent with the kex values for MN19 decreasing in both stem 1 and at the ligand-binding site. Based on the temperature dependence of the kex values, we find that MN19 is more dynamic than MN4 in the free and both ligand-bound forms. For MN4, ligand-binding results in the reduction of dynamics that are localized to the binding site. These results demonstrate that an aptamer in which the base pairs are preformed also experiences a reduction in dynamics with ligand binding. |
| Databáze: | OpenAIRE |
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