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2008 - Effect of Clotrimazole on the Pump Cycle of the Na,K-ATPase

Icon N1   SURFE²R ONE (a predecessor model of SURFE²R N1) publication in Biophysical Journal (2008)

Authors:
Bartolommei G., Devaux N., Tadini-Buoninsegni F., Moncelli M., Apell H.-J.

Journal:
Nature Scientific Reports (2008) 95(4): 1813–1825


Abstract:

The effect of the antimycotic drug clotrimazole (CLT) on the Na,K-ATPase was investigated using fluorescence and electrical measurements. The results obtained by steady-state fluorescence experiments with the electrochromic styryl dye RH421 were combined with those achieved by a pre-steady-state method based on fast solution exchange on a solid supported membrane that adsorbs the protein. Both techniques are suitable for monitoring the electrogenic steps of the pump cycle and are in general complementary, yielding distinct kinetic information. The experiments show clearly that CLT affects specific partial reactions of the pump cycle of the Na,K-ATPase with an affinity in the low micromolar range and in a reversible manner. All results can be consistently explained by proposing the CLT-promoted formation of an ion-occluded-CLT-bound conformational E2 state E2CLT(X2), that acts as a “dead-end” side track of the pump cycle, where X stands for H+ or K+. Na+ binding, enzyme phosphorylation, and Na+ transport were not affected by CLT, and at high CLT concentrations ~1/3 of the enzyme remained active in the physiological transport mode. The presence of Na+ and K+ destabilized the inactivated form of the Na,K-ATPase.


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