The static head method for determining the charge stoichiometry of coupled transport systems Applications to the sodium-coupled d-glucose transporters of the renal proximal tubule |
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Authors: | Yoshifumi Fukuhara RJames Turner |
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Institution: | Laboratory of Kidney and Electrolyte Metabolism, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20205 U.S.A. |
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Abstract: | The static head method for determining the charge stoichiometry (the number of moles of charge translocated per mole of substrate) of a coupled transport system is presented. The method involves establishing experimental conditions under which a membrane potential exactly balances the thermodynamic driving force of a known substrate gradient. The charge stoichiometry can then be calculated from thermodynamic principles. In contrast to the usual steady-state method for determining charge stoichiometry in cell suspensions and vesicle preparations, the static head method is applicable to systems which are not capable of maintaining a constant membrane potential over time. The charge stoichiometries of two renal sodium coupled d-glucose transporters previously identified in brush-border membrane vesicle preparations from the outer cortex (early proximal tubule) and outer medulla (late proximal tubule) are determined. The charge stoichiometries of these transporters are in good agreement with their sodium/glucose coupling ratios arguing against the possibility that glucose transport is coupled to ions other than sodium in these membranes. |
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Keywords: | Chemiosmotic theory Charge stoichiometry Coupled transport Glucose transport Static head method (rabbit kidney brush-border membrane) 10 mM Tris-Hepes 10 mM Hepes (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) buffered with Tris to pH 7 4 |
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