| Autor: |
Erika, Aloi, Jin-Hua, Guo, Rita, Guzzi, Ren-Wang, Jiang, Lucy Kate, Ladefoged, Derek, Marsh, Mikael, Esmann, Rosa, Bartucci, Natalya U, Fedosova |
| Rok vydání: |
2020 |
| Předmět: |
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| Zdroj: |
Biophysical journal. 120(13) |
| ISSN: |
1542-0086 |
| Popis: |
Spin labels based on cinobufagin, a specific inhibitor of the Na,K-ATPase, have proved valuable tools to characterize the binding site of cardiotonic steroids (CTSs), which also constitutes the extracellular cation pathway. Because existing literature suggests variations in the physiological responses caused by binding of different CTSs, we extended the original set of spin-labeled inhibitors to the more potent bufalin derivatives. Positioning of the spin labels within the Na,K-ATPase site was defined and visualized by molecular docking. Although the original cinobufagin labels exhibited lower affinity, continuous-wave electron paramagnetic resonance spectra of spin-labeled bufalins and cinobufagins revealed a high degree of pairwise similarity, implying that these two types of CTS bind in the same way. Further analysis of the spectral lineshapes of bound spin labels was performed with emphasis on their structure (PROXYL vs. TEMPO), as well as length and rigidity of the linkers. For comparable structures, the dynamic flexibility increased in parallel with linker length, with the longest linker placing the spin label at the entrance to the binding site. Temperature-related changes in spectral lineshapes indicate that six-membered nitroxide rings undergo boat-chair transitions, showing that the binding-site cross section can accommodate the accompanying changes in methyl-group orientation. D |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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