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Ouabain and marinobufagenin binding induce different conformations of Na,K-ATPaseтезисы доклада
- Авторы: Klimanova E.A., Petrushanko I.Y., Mitkevich V.A., Anashkina A.A., Orlov S.N., Lopina O.D., Makarov A.A.
- Сборник: Special Issue: 40th FEBS Congress
- Серия: The Bioochemical Basis Of Life
- Том: 282
- Тезисы
- Год издания: 2015
- Первая страница: 224
- Последняя страница: 224
- Аннотация: Na,K-ATPase, an ubiquitous ion pump, provides active transport of Na+ and K+ across plasma membrane in all types of animal cells. Moreover Na,K-ATPase is a receptor selectively responding to the changes in endogenous CTS level. Ouabain and marino- bufagenin are cardiotonic steroids (CTS), which specifically inhi- bit Na,K-ATPase activity. These CTS inhibit transport function of Na,K-ATPase in cells in the same concentrations, however they induce cell death characterized by different values of IC50. The reason for their different physiological effect is still not clear. Applying fluorescence measurements, isothermal titration calo- rimetry (ITC) and molecular modelling we have shown that bind- ing of ouabain and marinobufagenin cause different structural changes in Na,K-ATPase. Using fluorescence labeling we have shown that binding of both CTS with Na,K-ATPase shift confor- mation of Na,K-ATPase closer to the E1-state, and marinobuf- agenin induces more significant response than ouabain. Ouabain and marinobufagenin inhibit Na,K-ATPase hydrolytic activity with similar IC50 value (~1lM). However using ITC we observed a 17-fold higher affinity for binding of Na,K-ATPase in the E2P state to ouabain compared to marinobufagenin. The binding of ouabain to the enzyme is enthalpy-driven. In contrast, marino- bufagenin binding has a reduced enthalpic contribution and a larger entropic component. According to the ITC data both CTS bind to the same site. Molecular modeling predicts that ouabain is located deeper inside the binding site than marinobufagenin. According to our data ouabain and marinobufegenine may induce different conformations of Na,K-ATPase which support- ing binding of different proteins to the enzyme. Supported by the Russian Scientific Foundation (grant #14-14-01152).
- Добавил в систему: Климанова Елизавета Андреевна