Conformational changes of Na,K-ATPase probed with eosin Y.
Time-resolved fluorescence and binding studies have been carried out on Na,K-ATPase in the presence of the fluorescent dye eosin Y to obtain thermodynamic and kinetic parameters for the interaction of the enzyme with different cations. Eosin Y binding is indicated by a 3 ns fluorescence decay process and is observed only in the presence of mono- and divalent cations. This type of cation binding is interpreted as a nonselective electrostatic interaction, with negatively charged groups of the enzyme providing a high-affinity eosin Y binding site. Eosin Y binding is observed only under conditions where the enzyme exists in the conformational state F1. The kinetic parameters of eosin Y binding have been determined employing stopped-flow fluorometry.
Max-Planck-Institute of Biophysics, Kennedy-Allee 70, D-60596, Frankfurt, Germany.
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Lewitzki
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Schick
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Hutterer
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Schneider
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Grell