Oligomycin interaction with Na,K-ATPase: oligomycin binding and dissociation are slow processes.
Oligomycin interacts with the Na,K-ATPase by increasing the apparent Na+ affinity in the non-phosphorylated state of the enzyme. This property is used to estimate rate constants attributed to oligomycin binding and dissociation reactions with Na,K-ATPase. The rate constants are determined indirectly, employing stop-flow fluorimetry of eosin, the fluorescence of which is a marker for the E1 state of the enzyme, i.e. for Na+ binding. The second-order rate constants derived for oligomycin binding are in the range (6-12).10(4) M-1 s-1 at 6 degrees C for both shark rectal gland and pig kidney enzyme. Rate constants for dissociation of the enzyme-oligomycin complex are about 0.05 s-1 at 6 degrees C. The slow rates of binding and dissociation suggest that oligomycin acts from within the membrane lipid phase rather than from the aqueous phase. The dissociation constant at 6 degrees C for the enzyme-oligomycin complex can be calculated to be about 1 microM for shark enzyme and about 2 microM for kidney enzyme, at pH 7.0 in 2 mM NaCl.
Esmann
M
M
Institute of Biophysics, University of Aarhus, Denmark.
D000818:Animals; D004801:Eosine Yellowish-(YS); D007700:Kinetics; D008563:Membrane Lipids / Q000378:metabolism; D009840:Oligomycins / Q000378:metabolism*; D012754:Sharks; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D013050:Spectrometry, Fluorescence / Q000379:methods; D013552:Swine
D008563:Membrane Lipids; D009840:Oligomycins; D000254:Sodium-Potassium-Exchanging ATPase; D004801:Eosine Yellowish-(YS)